Understanding Science, Biology, and Physics

TTAG On Hiatus

2011-09-17T13:53:00.000-07:00

We're currently fixing things up at the moment.

Meanwhile, here's an article on how why light eyes may be more attractive. Limbal rings on light eyes are more noticeable, by the way.

TTAG Summary: The transcriptional program of sporulation in budding yeast.

2011-09-14T18:00:00.000-07:00

S. Chu, J. DeRisi, M. Eisen, J. Mulholland, D. Botstein, P. O. Brown, I. Herskowitz. 1998. The Transcriptional Program of Sporulation in Budding Yeast. Science, 282, 699-705.

Science 1998 Nov 20;282(5393):1421.

Here's my review on the paper.

So under starvation conditions, diploid Saccharomyces cerevisiae produce stress-resistant haploid spores in a process called sporulation. In order to successfully sporulate, diploid cells must undergo meiosis to reduce the ploidy of cells, and spore morphogenesis to package the genome in a prospore membrane, then the protective spore wall. The complexity of sporulation requires regulated changes in gene expression in order for the stages to progress in a coordinated, sequential manner.

The authors of the article are investigating every gene involved in sporulation and how different regulatory sites will affect when these genes are induced. The authors intend to categorize these genes based on their time of induction then link each category’s unique temporal profile to changes within the cell during the course of sporulation. Thus, they are able to hypothesize the functions of genes with previously unknown functions based on the time of their induction during sporulation.

To induce sporulation, cells were put on nitrogen-deficient medium. Sporulation is initiated by deficiencies in nitrogen and fermentable carbon sources, inducing a signal transduction pathway and arresting the cell at G1. At 7 different time points after sporulation initiation, they did a microarray analysis of 97% of all yeast genes to measure the changing abundance of mRNA transcripts of each gene over time compared to vegetative cells. The 7 different time points were based on the time of induction of DMC1, SPS1, DIT1, and SPS100, which represent the expression patterns for early, middle, mid-late, and late genes, respectively. Northern and microarray analysis were used to confirm the time of induction and relative transcript abundance of each of the four genes. Results from the microarray showed that, over the course of sporulation, 500 genes were induced and about 500 were repressed. The induced genes were categorized into seven groups (Metabolic, Early I, Early II, Early Middle, Middle, Late-Middle, and Late) based on the time of initial induction.

The metabolic group is induced first, and functions to acclimatize the cells to nitrogen-starved conditions. They have different temporal expression profiles, despite having the same URS1 sequence, and so must be regulated by other means. The majority of genes with the early I pattern have a URS1 site or a core URS1 site. They are induced after .5 hours and play a role in synapsis and homologous recombination. Genes with the early II pattern are less likely than early I genes to have putative URS1 sites, which may explain why their induction is slightly delayed. Early-middle genes are induced at 2 hours and continue to increase even after7 hours. Some genes that display this pattern have a role in spindle pole body dynamics. Middle genes, induced from 2-5 hours, are regulated primarily by Ndt80. Ectopic expression induced many genes with an MSE site in vegetative cells. Genes in vegetative cells that remain uninduced are likely to require additional factors. In addition to expressing Ndt80 in non-sporulating cells, the authors inactivated Ndt80 expression in sporulating cells. Many genes were expressed at a third the level of wild-type cells, resulting in arrest during prophase. However, other genes, induced independently of Ndt80, required additional input to be induced. The mid-late class of genes is induced after 5-7 hours, and the genes usually have an NRE (negative regulatory element) site in addition to the MSE site that delays their induction compared to middle genes. This class includes genes that contribute to prospore membrane formation. The late class of genes is induced after 7 hours and includes genes involved in spore maturation. Each class of genes has a common regulatory sequence in the promoter region targeted by a specific transcription factor. Binding of the transcription factor to its target site induces the gene’s transcription. Ume6/Ime1 recognizes the URS1 (Ume cognate cis-acting regulatory sequence) found upstream of early genes. Genes with MSE sites (middle gene sporulation element) induced by Ndt80 protein. Proteins that induce mid-late and late genes have not been discovered at the time of publication. Temporal expression patterns during sporulation are possible due to sequences such as URS1 and MSE, allowing induction of an entire set of genes at different stages of meiosis and spore morphogenesis when they are required. By linking what a gene does to how it is regulated by different transcription factors, the authors are able describe the general program of sporulation.

They needed to show that gene induction of the different classes was happened concurrently with physiological changes in cell, so the cells were assessed cytologically. At various time points, DAPI (4’-6’diaminidino-2-phenylindole) was used to stain the nuclei of sporulating cells to determine how many nuclei were present in the cells and measure the rate of meiosis. At around 5 hours, after the middle genes have been induced, mononucleate cells become binucleate. By 9 hours, tetranucleate cells outnumber mononucleate cells. Electron microscopy was used to determine the progress of spore maturation. From 5 to 9 hours, the rate of spore formation rose dramatically: from 0 percent to under 40. By 11.5 hours, the percent of mature spores had risen to equal that of immature spores. These results support the role of mid-late genes in prospore membrane formation and late genes in spore maturation because the cytological changes coincided with the gene’s induction.

The authors’ results show that in sporulation the time of induction of a gene is strongly correlated to its function and the mechanism of regulation. This correlation makes genes with unknown functions candidates for different processes based on their time of induction. The authors use this correlation as part of their strategy to propose the roles many genes such as Spo69, Spo70, and Spo71. The authors also compare homologous proteins with known functions in Drosophila and Xenopus and Caenorhabditis elegans to Spo70, a sporulation-specific protein, to determine whether the protein had the same function in yeast. In conclusion, the authors used temporal induction patterns of every gene in S. cerevisiae to propose gene functions for many previously uncharacterized genes. Genes each class had characteristic regulatory sequences that determined the time of induction. Time of induction during sporulation is tied with gene function as transcripts are induced when they have a role to play in the specific stages of the program. Thus, regulatory sites such as MSE and URS1 can be used to determine gene function.

Question of the Day: Liquid Mechanics Involving Protein Powder

2011-09-13T18:53:00.000-07:00

WHY DOESN'T THIS EVER MIX WELL IN WATER?!?!

Headache01 asks "Why doesn't powder mixed into drinks (like whey protein powders or cocoa) dissolve very well? It clumps into balls that are wet on the outside but remain dry on the inside. How can I make my whey mix better?"

Well, given that proteins, and especially mixture of proteins, are amphiphilic (meaning the molecule has a part with an affinity for water, and another for fat/oils), they tend to orient themselves into microscopic structures known as micelles. Micelles allow the molecules to isolate their lipophilic ends away from the water and their hydrophilic end towards water, ultimately forming a spherical, tubular, or sheetlike structure.

While micelles are a microscopic phenomena, a similar thing is what causes the clumps in your protein drink; along the protein powder-water interface there will be two-layer sheet type structures where the hydrophilic parts of the molecules are facing the water and the lipophilic parts stay away from it.

Wonder why you should avoid warm water when mixing powders? The heat causes the proteins to lose their secondary structure and become entangled with one another, making it difficult to break up the clump since the clump's outer surface has essentially polymerized. Thus, using cold water keeps the proteins tightly coiled and less likely to get entangled with each other.

I almost forgot! The solution is to wet each of the solid particles individually first before dispersing them (e.g. mix in a small amount of water to form a paste). This will ensure that they disperse well.

DNA Repair Systems

2011-09-12T21:41:00.000-07:00

During replication, DNA polymerase proofreads the newly synthesized strand, and improperly incorporated bases are removed by its 3' to 5’ exonuclease activity. In addition to proofreading, replication errors are corrected by the mismatch repair system. Mismatched bases change the conformation of the helix. In the mismatch repair system, the distortion is recognized and the region around the newly synthesized strand is excised. Some bacteria use methylase to differentiate between the old and newly synthesized strand. DNA polymerase then fills in the gap, using the older strand as a template. Other global systems commonly repair DNA in cells: base excision, photolyase, and nucleotide excision repair. In base excision repair, DNA glycosylase recognizes specific faulty bases, and hydrolyzes the glycosidic bond between the base and the deoxyribose backbone. AP endo/exonuclease then excises the single deoxyribose, and DNA polymerase fills in the gap. In photoreactivation, DNA photolyase recognizes and binds to thymine dimers, which cause a conformational change in the DNA helix. When light activates this enzyme, it breaks the covalent bonds of the ring, reversing UV damage. The nucleotide excision repair, helicases melt the duplex at the site of distortions, and a 12-13 residue long single-stranded DNA segment at this site is excised. DNA polymerase then fills in the gap left behind.

DNA damage can involve more than just wrongly incorporated bases. When double strand breaks or gaps occur, they are repaired either by non-homologous end joining (before replication) or homologous recombination (after replication, when sister chromatids can provide a template). In NHEJ, an exonuclease process the single stranded ends of the broken DNA, and ligases then directly join them together. The digestion of ends may result in the loss of nucleotides and mutations. Homologous recombination is less mutagenic because sister chromatids or other homologous regions are used as a template for repairing the gap.

Some mutations are induced as part of a cell’s response to stress and DNA damage. The presence of ssDNA induces the SOS response in E. coli. RecA binds to ssDNA filaments and ATP, activating RecA. Active RecA induces self-cleavage of the repressor of din genes, LexA, and the din genes, including the umuDC operon, are transcribed. The umuD and umuC gene products form a heterotrimer UmuD’(2)C, DNA Pol V, which will synthesize DNA across lesions irrespective of the residues on the template strand. For example, DNA Pol V may insert guanines opposite a thymine dimer or a cytosine opposite an apurinic site, something DNA Pol III cannot do. Thus, the SOS response and DNA Pol V allow the cell to continue replication of its genome despite DNA damage. Additionally, DNA Pol V’s mutagenic nature allows cells to mutate specifically when they are maladapted to their environment.

Mutagens and Mutagenesis

2011-09-11T21:29:00.000-07:00

Negatively charged DNA coiled around histones. Credit: Thomas Splettstoesser (from PDB 1EQZ)

Mutations are permanent changes in the genetic material passed down from one generation to the next. In order for the continuation of the species, each generation of an organism must faithfully replicate their genomes. In order for the species to evolve and adapt, some mutations must be allowed to occur. As the substrate of evolution, mutations are especially crucial for the maintenance of genetic variation in microbial populations, which do not rely on sex or meiosis.

In the laboratory, mutagens allow geneticists to study loss-of-function mutations and elucidate gene function. For instance, the function of the gene products involved in DNA repair was elucidated by introducing mutations in their respective genes and comparing mutants’ sensitivity to chemical and ionizing mutagen to the sensitivity of the wild-type organism.

Nitrous acid. Don't drink it.

For example, mitomycin C and 1,2,7,8-diepoxyoctane cause deletions while nitrous acid causes oxidative deamination of adenine and thymine to hypoxanthine and xanthine, respectively. Diepoxybutane affects G:C base pairing. N-methyl-N-nitrosourea and N-methyl-N’-nitro-N-nitrosoguanidine methylate bases, causing base-pair substitutions and baseless deoxyriboses. Ethidium bromide has a ring structure that intercalates bases and stretches the duplex, leading to frameshift mutations. Incorporation of base analogs, with higher rates of tautomerization, into DNA during replication will also lead to mutations. Ultraviolet irradiation is also mutagenic, and comes in three forms (in order of increasing energy) : UVA, UVB, UVC. UVC is the most lethal to bacteria, but all of them will induce the formation of thymine dimers. UV formation of a cyclobutane ring between the two adjacent thymines, called a thymine dimer. Without chemical and ionizing mutagens, the rate of mutagenesis would be so low, mutant yield would be low in the laboratory, and isolating them would be impractical.

Spontaneous mutations can occur when bases react with water or other natural species in the cellular environment. For example, cytosine deaminates to uracil, methylcytosine deaminates to thymine, adenine . Bases can tautomerize, switching from keto to enol form. Each form has a different base pairing property. For example, a guanosine in its keto form base pairs with cytosine, but a guanosine in its enol form will base pair with thymine. Thus, a switch from the keto to the enol form results in a G:C to A:T transition in one of the daughter cells. Mutagenic base analogs like 5-bromouracil tautomerize and cause transitions this way. However, DNA damage can involve more than just incorrectly incorporated bases. When double strand breaks or gaps occur, they are repaired either by non-homologous end joining (before replication) or homologous recombination (after replication, when sister chromatids can provide a template). In NHEJ, an exonuclease process the single stranded ends of the broken DNA, and ligases then directly join them together. The digestion of ends may result in the loss of nucleotides and mutations. Homologous recombination is less mutagenic because sister chromatids or other homologous regions are used as a template for repairing the gap.

Given that these changes to the DNA do not result in mutations unless they are fixed after a round of replication, the cell must repair the DNA before replication ends. This Monday night, we'll go over how the cell detects aberrations and repairs them!

Hubble Telescope Discovered Dark Matter

2011-09-10T02:10:00.000-07:00

Dark matter makes up about 73% of our universe, according to space.com.

Well, a discovery the Hubble Telescope was responsible for is dark matter, which has a unique composition compared to gas and dust in galaxies, and makes up most of the universe. A dark matter ring was discovered in 2007 at the site of two galaxy clusters’ collision, 5 billion light-years away from Earth. It was detected by the deflection, or bending of light in background galaxies when looking at the two galaxies’ collisions. The dark matter is probably a result of the high gravity following the collision. Evidence for dark matter, therefore, is indirect, only detectable when galaxies in the foreground caused distortions in the light of galaxies in the background.

Currently, the Hubble Space Telescope’s ACS is being used to survey dwarf galaxies 250 million light years away, in the Perseus Cluster. It is still too far away to detect small stars that may be too faint, but larger, more luminous stars are still readily detected. The Hubble Space Telescope revealed that at least 12 of the stars in the Perseus Cluster require dark matter. The stars appear smoother, scientists believe, because of the presence of dark matter, relative to the stars in spiral galaxies such as the Milky Way. The dark matter would protect these dwarf galaxies, whose stars have a high mass to light ratio (Hubble made it possible to calculate their mass by measuring the amount of X-ray radiation emitted by the hot gases of the stars), from being destroyed by outside gravitational pull. The discovery of this phenomenon would have been impossible without the incredibly high detail provided by a space telescope such as Hubble. As previously mentioned, ground telescopes have too much background light interference and not enough resolution for study of individual stars in galaxies outside our own. [1] [2]

1. Hubble Finds Ring of Dark Matter. 5 Oct 2010. Available:

http://www.spacetelescope.org/news/heic0709/

2. Hubble Provides New Evidence for Dark Matter in Small Galaxies. 14 Nov 2010. Available:

http://hubblesite.org/newscenter/archive/releases/2009/11/full/

Edit: If there's anything I want you to get from these recent posts on the Hubble, it's that funding for space programs helps us understand so many new things about the universe we live in and its beautiful physical laws. Dark matter would have probably remained a mystery if it had not been for the images from Hubble. Please write to your representatives urging them to continue supporting NASA or PBS, which has fascinating programs to educate us all.

Cepheids

2011-09-07T17:39:00.000-07:00

The smaller of the two stars is a Cepheid variable. Credit: ESO/L. Calçada

Charged couple devices on the Hubble are what helped us figure out the rate of expansion in the universe by using the opacity of stars near us. Dense stars have no temperature gradient, while less dense stars do. NICMOS and the infrared sensors were used to detect any temperature gradients in stars to determine their relative density. It detected stars that were variable between a dense and a less dense state, with observable changes in pressure and temperature. These stars are called Cepheids. This variation’s period was shown to be correlated with the apparent luminosity of the star, which increases and decreases at a regular interval.

The star’s relative distance can be determined now by detecting the change in the pulsating luminosity over time. Stars very far away are dimmer, and stars closer to us appear brighter. The scientists measured the differences between the brightness of different galaxies, cross-checked this data with other distance indicators, and determined how fast these galaxies are moving apart from each other. Cross-checking is necessary because other factors can affect the luminosity of stars, and having only one variable to determine the distance between stars would lead to many inaccuracies. For example, big stars are very dusty, but also very short-lived, which was a limitation for scientists, and gravity of different galaxies can affect the rate of movement, distorting the rate of expansion of the universe. It is possible for us to figure out the rate of expansion because the space telescope can adjust for the relative motion caused by the pull of gravity of galaxies nearby. It was a big step in the attempt to determine the rate of the expansion of the universe

Source:

WMP, the Expansion of the Universe. 12 Nov 2010. Available: http://map.gsfc.nasa.gov/universe/uni_expansion.html

TTAG In-Depth Look: Instrumentation on the Hubble Space Telescope

2011-09-06T20:15:00.000-07:00

http://arstechnica.com/science/news/2010/04/hubble-turns-20-a-retrospective-in-pictures.ars (click to enlarge)

The Hubble Space Telescope is responsible for many discoveries that revolutionized our view of the universe. A few observations here and there of phenomena in a minute portion of the sky is all that is needed to infer how our entire universe works. To illustrate, a few of Hubble's images lead to the discovery of dark matter. Yes, physics is that elegant.

Since its launch in 1990, the Hubble Space Telescope has provided the whole world with a detailed view of the universe that was never before possible. Given that the Hubble Space Telescope is outside of the Earth’s atmosphere, the images are clearer and sharper than any telescope located on the Earth’s surface. This is because the light that the ground telescope receives goes through air first. The molecules in the air distort and diffract light, and they absorb certain wavelengths of light, such as ultraviolet and gamma radiation. Space telescopes don’t have this problem outside of the atmosphere. Since it is in orbit above Earth’s atmosphere, there is no background light from Earth, and light is unaltered and undistorted by the molecular gases in our atmosphere.

The Hubble Space Telescope was launched into low Earth orbit by the space shuttle Discovery on April 25th 1990, It travels about 5 miles per second, takes 97 minutes to complete an orbit around Earth. [15] Even though it was meant to have been launched in 1983, it was delayed seven years because of financial constrains, and a cautious Congress and populace after the Challenger disaster a few years prior in 1989.

It is expected to re-enter into Earth’s orbit within the next few decades, due to orbital decay and drag, at which time a new space telescope is planned to replace it. Retrieval is considered not practical as it would risk the lives of people and would be too costly to do, as the shuttle program is already retired.

Instrumentation

The Hubble Space Telescope works to collect light from a wide spectrum and focus it, allowing us to see farther and clearer than the human eye possibly could. It is a Ritchey-Cretien Cassegrain reflector, meaning incoming light that travels from distant stars will come and bounce off a primary concave mirror and hit a secondary convex mirror, which focuses light through a small hole in the center of the primary mirror. The primary mirror’s diameter and size is one of the main determinants for how much light the telescope can collect, and compared to the large ground telescopes in observatory towers, the one on the Hubble would be considered small, at just 2.4 meters. [15] After incoming light passes through the hole in the primary mirror, it can be collected by the instrumentation on the Hubble Space Telescope. The mirrors must be very smooth, uniform don to 1/800,000th of an inch, and are treated with various coats to increase transmissibility of light, and to protect the mirror itself from warping due to solarization and aging due to ultraviolet light. The complex structure of the two mirrors, apertures, and trusses that support it is called the Optical Telescope Assembly, or OTA. [16]

Even small aberrations of the mirrors can cripple the Hubble Space Telescope. For example, after its launch, they discovered that the main mirror’s curvature had problems, but that was fixed after the first service mission in 1993. Since its launch, the Hubble Space Telescope’s fine instrumentation have required five different service missions: Service Mission 1 in 1993, Service Mission 2 in 1997, Service Mission 3A in 1999 to put in a new computer and gyroscopes, 3B in 2002 to replace the FOC with the Advanced Camera for Surveys, repair the NICMOS Cryocooler, and the most recent one, 4, in May 2009 to repair the Advanced Camera for Surveys, the Space Telescope Imaging Spectrograph, replace the Fine Guidance Sensoring System and two gyroscopes. [2][5][6][7][8] To maintain accuracy and for the data retrieved by the telescope be useful for scientific observation, constant upkeep and calibrations of the instruments must be made. The instruments themselves will now be described.

NICMOS and Cryocooler

NICMOS, or Near Infrared Camera and Multi-Object Spectrometer was installed on the Hubble Space Telescope in February of 1997, during the Servicing Mission 2. In short, it seeks out heat in the form of infrared radiation.

It is composed of three cameras, each with their own field of view, with 256x256 HgCdTe Rockwell sensor, sensitive to wavelengths of .8 micrometers to 2.5 micrometers. They are housed in a cryogenic dewar, maintaining a constant working temperature between 58 and 60 degrees Kelvin. The dewar is composed of three shields to keep NICMOS cool: the VCS, or Vapor Cooled Shield, the Thermo-Electric Cooled Inner Shield, and the Thermo-Electric Cooled Outer Shield. The hybrid nitrogen and aluminum dewar was supposed to keep it at just 58 degrees Kelvin, but a gap in the Vapor Cooled Shield resulted in an unanticipated heat load. Fluctuations in temperature where the cameras are housed means the cameras will need to calibrate more often.

This is where the Cryogenic Cooler comes into play, maintaining temperature stability within .5 degrees Kelvin. It circulates neon gas through a cooling loop, using high-speed centrifugal machines that do 7000 revolutions a second to compress gas, removing heat from the gas. [18]

NICMOS is useful because it captures information about infrared light, which reaches Earth from very far away, unaffected by interstellar dust, unlike visible light.

Advanced Camera for Surveying

The Hubble Telescope’s FOC was replaced by the ACS in 2002, during the Service Mission 3B. It has three cameras that are sensitive to a wide spectrum of light, from ultraviolet to near infrared (wavelengths of 1,200 to 10,000 angstroms), and since it has high contrast even near bright stars, it can be used to study galaxies and black holes very far away, where light from the ancient universe has just arrived at Earth. ACS has many components that make it versatile and useful for scientific observations. For instance, ACS is sensitive to ultraviolet light because it has a solar blind camera, or SBC, to block out visible light.

The Wide Field Channel of the ACS has two cameras with a resolution of 2048x4096 pixels each (for a total of 4k by 8k pixels), and its wide view frame is used to survey galaxies and the positions of stars within those galaxies. It is responsible for the Ultra Deep Field images of the universe.

The High Resolution Channel of the ACS has one camera with a resolution of 1024x1024, and is used primarily to detect ultraviolet light. The HRC has a coronagraph component that increases the contrast near stars tenfold. The High Resolution Channel allows us to “zoom” in, with a smaller field of view, but the images have a greater angular resolution. While the WFC and HRC are mainly on the red and blue regions of light, there’s a Multi-Anode Microchannel Array has no electronic noise and is sensitive to ultraviolet light, but not visible light. The images are 1k by 1k pixels, which is less than the WFC and HRC. Only the Solar Blind Camera is working right now. [10]

COS, STIS, and Ultraviolet Light

Spectrographs allows us to see precise information about incoming light that would otherwise be very faint and unusable. Incoming light is broken down to different wavelengths of light, and the amounts of each light at specific wavelengths are plotted on a graph. The light would have a unique fingerprint of different wavelengths, which can be studied.

COS, or the Cosmic Origins Spectrograph, looks at specific stars.

STIS, or Space Telescope Imaging Spectrograph, primarily has a larger field of view, focusing mainly on galaxies or a system of stars.

The Hubble Space Telescopes main mirrors are coated with magnesium fluoride 1 x 10-6 inches thick to improve the reflection of light at ultraviolet wavelengths. [16] Ultraviolet light, with a wavelength of 300 nanometers to 10 nanometers, is very high energy, so a special material that had a high energy gap and was resistant to absorbing the high energy from the photons was required. Magnesium fluoride not only helps maintain the integrity of ultraviolet light, the resistance to absorbing photon energy prevents warping of the mirrors due to solarization. [17][19]

Pointing and Guidance Systems

A system of fine guidance sensors, magnometers, solar sensors, and gyroscopes help keep the telescope position and point itself to collect light and data from specific areas in our universe. Since the Hubble Space Telescope is constantly moving in low orbit at a rate about 5 miles per second, a wealth of stabilizing components is needed to ensure the telescope is pointed at one spot for sufficient periods of time. The position and orientation of the telescope is also crucial because solar energy needs to hit the panels to power the telescope, and to keep the Sun’s heat from hitting only one side of the telescope. [19][4]

Currently, 3 gas-bearing gyroscopes rotate up to 19,000 rpm help stabilize the telescope, and with the help of up to three fine guidance sensors, the Hubble Space Telescope can help determine star locations that are 10 times more precise than a ground telescope. The guidance system also helps determine the position of stars, the distances between them, and the distance scale of the universe. Magnometers and CSS also help determine the telescope’s position relative to Earth’s magnetic field and the sun, respectively. Magnetic torquers and reaction wheel actuators help “lock” the telescope on to a planet or point in space. [4][19]

Design for Power

The Hubble Space Telescope is powered by 57 kilograms of nickel-hydrogen batteries when solar power is not available for 36 minutes in the Earth’s shadow. [3] Each orbit is 97 minutes, so it spends roughly 37 percent of its time without solar energy, so batteries are important. They must be able to power and sustain the telescope when solar power isn’t available. There are two modules consisting of a total of six batteries. Each module weighs 460 pounds and measures 36 inches long, 32 inches wide, and 11 inches high. There are three batteries per module, with each battery having 22 individual nickel-hydrogen cells placed in series.

Even though each battery has a total of 88 amp-hrs of capacity, on the Hubble Space Telescope, the practical maximum is 77 amp-hrs due to a limitation with heat dissipation. With a total of six batteries, the total energy supply becomes 450 amp-hrs. The batteries have lasted almost 13 years more than originally planned, 18 total. The ones that will replace them will be even better, built with the “wet slurry” process over the dry method. The “wet slurry” process allows for better porosity of the solid metallic powder over the dry method, in which the metallic powder is simply impacted into the battery cells. [3]

With the new batteries, the Telescope be used for normal scientific purposes on battery power for nearly 5 orbits, which is nearly 7.5 hours of operation, assuming the batteries are fully charged. [14] This means power for the Hubble Space Telescope is supplied redundantly in the sunlight, so if one source of power fails, the telescope will still be in working condition.

When the Hubble Space Telescope is within the day time portion of its orbit, about 61 minutes of the 97 minute orbit, it uses an array of four solar panels to power the telescope and to charge the batteries. They were replaced in the Servicing Mission 3B in 2002 with smaller, less flexible panels that produced less than a third more power than the pre-existing ones. Solar arrays supply 5k watts of energy to power the Hubble Space Telescope. This extra power supply makes running more instruments simultaneously on the telescope possible, so more institutions and scientists can use the telescope at the same time. [13]

1. Hubble Space Telescope Servicing. 5 Oct 2010. Available:

http://www.nasa.gov/mission_pages/hubble/servicing/SM4/main/FGS_FS_HTML.html

2. Servicing Misson 4. 5 Oct 2010. Available:

http://sm4.gsfc.nasa.gov/technology/sm4_acs.php

3. Servicing Mission 4 <Batteries>.

http://www.nasa.gov/mission_pages/hubble/servicing/SM4/main/Battery_FS_HTML.html

4. Servicing Misson 4 <Gyropscopes>. 5 Oct 2010. Available:

http://www.nasa.gov/mission_pages/hubble/servicing/SM4/main/Gyro_FS_HTML.html

5. Servicing Mission 3B. 5 Oct 2010. Available:

http://hubble.nasa.gov/missions/sm3b.php

6. Servicing Mission 3A. 5 Oct 2010. Available:

http://sm3a.gsfc.nasa.gov/overview.html

7. Servicing Mission 2. 5 Oct 2010. Available:

http://hubble.nasa.gov/missions/sm2.php

8. Servicing Mission 1. 5 Oct 2010. Available:

http://hubble.nasa.gov/missions/sm1.php

9. NASA Hubble. 5 Oct 2010. Available:

http://www.nasa.gov/mission_pages/hubble/spacecraft/index.html

10. The Hubble Program – Technology. 5 Oct 2010. Available:

http://hubble.nasa.gov/technology/instruments.php

11. Hubble Finds Ring of Dark Matter. 5 Oct 2010. Available:

http://www.spacetelescope.org/news/heic0709/

12. WMP, the Expansion of the Universe. 12 Nov 2010. Available: http://map.gsfc.nasa.gov/universe/uni_expansion.html

13. Hubble Servicing Missions. 25 Oct 2010. Available:

http://hubblesite.org/the_telescope/team_hubble/servicing_missions.php

14. Hubble the Telescope. 25 Oct 2010. Available:

http://hubblesite.org/the_telescope/nuts_.and._bolts/spacecraft_systems/#power

15. Hubble Telescope Essentials. 26 Oct 2010. Available:

http://hubblesite.org/the_telescope/hubble_essentials/

16. Optical Assembly. 12 Nov 2010. Available:

http://hubblesite.org/the_telescope/nuts_.and._bolts/optics/

17. Hubble Provides New Evidence for Dark Matter in Small Galaxies. 14 Nov 2010. Available:

http://hubblesite.org/newscenter/archive/releases/2009/11/full/

18. NICMOS. 12 Nov 2010. Available:

http://www.stsci.edu/hst/nicmos/documents/papers/2002-ncs_spie_paper1.pdf

19. Hubble – Fine Guidance Sensors. 5 Nov 2010. Available:

http://hubblesite.org/the_telescope/nuts_.and._bolts/instruments/fgs/index.php

Question of the Day: Occupational Hazards

2011-09-03T01:34:00.000-07:00

Question of the day for you folks: what kind of occupational hazards do you encounter at your job?

[Update: Deleted the post 9/10/11... just left the question!]

Assay for β-galactosidase Specific Activity

2011-09-01T00:08:00.001-07:00

As previously mentioned, β-galactosidase cleaves lactose into galactose and glucose. However, these products of the natural reaction are not easily quantifiable. That is why instead of lactose, we introduce ONPG to the cells. (ONPG, o-nitrophenyl-B-galactoside, is a substrate but not an inducer of the lac operon.) ONPG, colorless, is cleaved by β-galactosidase into galactose and orthonitrophenol, which is yellow, has a λmax of 420. The amount of o-nitrophenol can be quantitatively reported by looking at the change in the absorbance at 420 nm. The ΔA420/min is the total rate of the enzyme’s activity. To find the specific activity of β-galactosidase in relation to cell concentration, the rate is divided by the concentration of bacteria cells. Thus, the specific activity can be expressed by the following equation: ΔA420/min/OD600/ml. If the culture is not in log phase and in the death phase, dead cells contribute to the optical density but they can’t produce β-galactosidase to contribute to the total activity, leading to a deflated specific activity value for the enzyme.

Let's go it over step by step, shall we? I'll walk you through it.

A culture in the log phase is needed, as inducing gene expression works best during this phase. Inoculate liquid broth with E. coli and grow it at 37⁰ C. The broth shouldn’t have any glucose, as this would block any lacZ expression. Using the broth as a blank, transfer the culture into cuvettes and take the OD600 of the culture until it reads approximately .5-.6. In a tube with the culture, add the phosphate Z buffer, which keeps β-galactosidase working efficiently. Add a drop of sodium docecyl sulfate (SDS) and a drop of chloroform, which will lyse the cells, stop translation, but leave β-galactosidase intact and available for ONPG hydrolysis. Keep this in a room temperature water bath until the temperature has equilibrated. It should be stable for a while, preventing time errors. Add ONPG to each tube being assayed. This will be time 0, the start of the reaction. An Eppendorf tube with distilled water, Z buffer, and ONPG should be used as a control to make sure hydrolysis of ONPG is because of the enzyme, and not a spontaneous reaction in the mixture. If, after 5 minutes, the tubes turn yellow too quickly, the bacterial culture should be diluted and re-assayed, because that small window of time means small time errors will skew the results significantly. In addition, there needs to be an excess of ONPG during the assay, to ensure that the rate of hydrolysis is limited by, and therefore determined by, the enzyme concentration. If not enough ONPG is available, the increase in A420 would not correlate with increases in expression of galactosidase. The start of induction for this enzyme takes about 15-30 seconds, with a logarithmic increase when plotted against time, so sometimes reactions can go to completion is inappropriate amounts of ONPG are used. Use sodium carbonate (Na2CO3) once the yellow color is observed to stop the reaction. Na2CO3 inhibits β-galactosidase activity, thus stopping ONPG hydrolysis. Take the tubes and centrifuge them for 10-15 minutes at 5,000 g. Transfer the supernatant to cuvettes and measure the OD420. Visually confirm the readings are reasonable. A420 should be higher the darker the yellow. If the pellet and supernatant are not separated properly, the cells will scatter the light and invalidate the readings. During the entire reaction period, the temperature must be kept constant, because variable temperatures will affect enzyme activity and absorbance values.

The change in A420 can be converted to moles of ONGP converted to ONP+ by using Beer’s law and plugging in the molar extinction coefficient for ONP+, 4500/M/cm. The moles of ONP+ converted can be divided by the turnover rate of β-galactosidase to find out how much β-galactosidase is active.

The assay is used to determine relative expression of the lacZ gene and the specific activity of β-galactosidase, the lacZ gene product, in a bacterial culture. In uninduced cultures vs. fully induced cultures, there is a 1000% difference in β-galactosidase activity. Using this assay, the amount of time that it takes for the lac operon to become fully induced in a culture and be determined. Most importantly, we can manipulate the operon and insert a set of other structural genes downstream of the lac promoter by homologous recombination. Presence of a neoR cassette, too, allows us to select for these transgenic cells. Since the sequence will under the promoter’s regulation, the expression of the genes inserted is proportional to the expression of lacZ. Thus, we can indirectly determine expression of the inserted sequence by assaying B-galactosidase activity.

Love science? Write for TTAG!

2011-08-30T21:43:00.000-07:00

We're hiring!

The Truth About Genetics (TTAG) is looking for a few enthusiastic writers to join our team. If you're good, we'll pay you at least $20 per article you write! If you demonstrate a consistent schedule, you'll get a weekly sum.

Interested? Just submit an original sample of an article that either reviews a primary research article or explains a concept in biology or physics. Articles on TTAG can be either colloquial or formal, so the sample can be written in your own style. Either way, applicants must write authoritatively and clearly and demonstrate a passion for science.

Send your writing sample and introduce yourself to writers@thetruthaboutgenetics.com. We'll get back to you shortly.

Question of the Day: Physics or Genetics?

2011-08-29T20:54:00.000-07:00

Some people love physics and math. With a few observations here and there, a physicist can infer the whole workings of the universe. "Oh, the moons of Jupiter seem to orbit around Jupiter at various speeds." Ahah! "The speed of light is finite!" He can do this because physics is so elegant and its rules are so universal. Even though it's all theoretical, there's a sort of inevitability to physics that makes it so appealing to me. Reading Eintein and Hawking, I've come to discover how the universe works in one way, and that is how it has to be. Let me put it this way: the same principles governing the way an apple falls from a tree is also the same principle underlying black holes forming from pulsars billions of light years away. It's so fascinating.

On the other hand, genetics and biology don't follow universal rules, and theories involving such a dynamic and living thing are never absolute or inevitable. Life is so dynamic; a biologist cannot make accurate inferences about life billions of years ago based on life today, and we may never know. An organism has a gene for lactose catabolism? So many different ways it could have acquires those genes. Independently, through lateral gene transfer, through a million simultaneous base substitutions, etc. Not to belabor the point, but it is inelegant in contrast to physics.

So which do you prefer, biology or physics? Do the isotopes in the stars interest you more, or do you prefer the study of animals and the causes of diseases?

I mentioned isotopes because I needed something to segue to what I did today; I cleaned up a super radioactive microfuge in the lab I work at today. The readings of tritium were off the charts, and so I did a pre-cleaning liquid scintillation count and two post-cleaning counts. Given the inherent danger of exposure to radioactivity, I still managed to have a blast.

No Science Sunday: Ambien Restores Speech For An Hour

2011-08-28T01:40:00.000-07:00

A vibrant 24-year old man with cystic fibrosis suffers two massive strokes after headbutting a soccer ball. Doctors said the entire intellectual and visual area of his brain were essentially wiped out, and he would live his life as a vegetable. However, a medicine known as Ambien in the United States and Stilnox in Austrailia is able to restore his speech for an hour a day.

Their struggle, their remarkable spirit, and the bond Sally and Sam share is awesome. Awesome is definitely an overused word, but its use is appropriate here.

BRCA2 and Homologous Recombination

2011-08-27T16:04:00.000-07:00

BRCA2, Breast Cancer 2 susceptibility protein, is encoded by BRCA2 gene mapped to 13q12.3. BRCA2 repairs DNA damage through homologous recombinbation, the exchange of identical or nearly identical sequences of DNA. Homologous recombination occurs mostly during sexual reproduction or repair of DNA damage. Unrepaired, DNA damage leads to mutations in the cell line, and eventually, cancer. Without BRCA2 protein, our body's ability to repair DNA damage and avoid cancer are seriously hindered. That is why people with defective BRCA2 are susceptible to breast cancer. DNA damage goes unrepaired, and cells behave aberrantly.

Now, what's fascinating about BRCA2 is that we originally discovered its function by looking at its homolog in the fungi Ustilago maydis, the corn smut. Scientists sequenced the peptide chain the known damage repair protein in U. maydis and compared it with proteins of unknown function in humans. Proteins in humans that have the same function will have similar sequences, because amino acid sequence determines protein structure, affinity, and function. Amazing, isn't it?

People often ask me why I chose microbiology. Microbes are great organisms to study genetically because of their small and haploid genomes, short generation cycle, and their ability to uptake free DNA through transformation. Small genomes are easier to map. Being haploid organisms, they only have one set of DNA, so changes in the DNA will have corresponding changes in their phenotype. Being haploid, they're asexual, so bacteria and other prokaryotes (including some unicellular eukaryotes) produce genetic clones in subsequent generations. Microbes' ability to uptake free DNA helped geneticists to easily manipulate their genomes before the advent of gene targeting in mammals.

It was only after Dr. Mario Capecchi's research we use homologous recombination to edit specific genes in mammals and actually play around with higher animal's genetics.

Article by UCD on the function of homologous recombination proteins in different model organisms:

http://www.pnas.org/content/108/2/441.full.pdf

Every time they mention HR they mean homologous recombination.

Interested in gene targeting in mice? Find out more!

Video of Dr. Mario Capecchi, 2007 Nobel Laureate:

http://www.ibiomagazine.org/issues/march-2011-issue/mario-capecchi

Steps for homologous recbomination in mice:

http://www.bio.davidson.edu/courses/genomics/method/homolrecomb.html

An Overview of the Polymerase Chain Reaction

2011-08-25T19:08:00.000-07:00

The polymerase chain reaction (PCR) is a method that selectively replicates desired DNA segments in vitro. It requires that we design two primers, each 20-30 base pairs long, that will hybridize to the 3’ ends flanking the desired DNA segment. The forward primer and reverse primer are anti-parallel and complementary to the nonsense and sense strand, respectively. For instance, if the nonsense strand of the DNA has the sequence 5’ TTG CCA GAT 3’, the forward primer is 5’ ATC TGG CAA 3’. Additionally, restriction enzyme recognition sites can be added onto the 5’ end of these primers to facilitate ligation of the target DNA into a cloning vector.

First, these primers, along with Taq polymerase and other co-factors, are added to the target DNA that has been heated to 90°C for two minutes. The high temperature separates strands of the target DNA. Second, the sample is cooled to 42 to 65°C for 30 seconds, allowing the DNA will anneal with the primers. The temperature is raised to 72°C (the ideal temperature for polymerase) so Taq polymerase can extend the primers. 1 minute per 1kb of amplicon at 72°C.The thermal cycling is repeated for 25-30 times. Each cycle denatures the DNA, anneals the primers, and then extends the primers. The target DNA doubles in amount after every cycle. A temperature of 72°C is set for 5 minutes to ensure all primers have been fully extended.

Taq polymerase is a heat stable polymerase that remains active even after heating to 90°C. In addition to Taq polymerase, primers, and the target DNA, MgCl2, buffers, and 100-200 µM of each dNTP is added. Higher concentrations of dNTPs allow for faster cycles, but lower concentrations will increase fidelity. Taq polymerase has no 3’à5’ exonuclease activity, so having sufficient fidelity is important. Mg2+ is a required cofactor for polymerization, coordinating the dNTPs in the Taq’s active site.

To make sure the PCR went well and the primers hybridized correctly, we either sequence the amplicon directly or digest it with restriction enzymes and run it on a gel.

In nature, restriction enzymes protect bacteria from viral infection by cleaving foreign, DNA at specific recognition sequences. Bacteria have methyl transferases that methylate its genome, protecting it from cleavage. In lab, restriction enzymes are used to cut DNA sequence of interest. Treating DNA with a specific restriction enzymes and then running it on a gel allows us to see where recognition sites are located, and how frequent they appear in our sequence of interest.

EcoRV, for example, cuts blunt ends at 5’ GATATC 3’. If our PCR amplicon has 6 of these sequences, the amplicon will be cut 6 different times, resulting in 7 different sizes of DNA molecules. We can then separate the 7 different DNA fragments and determine their size with PCR. Different DNA molecules will contain a different number of recognition sites, so they will have unique fingerprints. The more kinds of restriction enzymes you use, the higher resolution the fingerprints will have. The fingerprints of your amplicon can then be compared to the fingerprint of the known sample. If they show the same bands, the PCR was successful.

Gel electrophoresis is a technique that separates DNA based on size and shape. DNA is loaded onto an agarose gel. A negatively charged cathode and a positively charged anode at the bottom of gel will apply an electric field to the gel. Because of phosphates in its backbone, DNA will migrate towards the anode at the foot of the gel. Larger or bulkier molecules of DNA will move slower through the gel matrix than smaller ones, which reach the foot of the gel more quickly. After a given time, the relative size of DNA molecules can be visualized by the distances they moved through the gel.

The gel is made with 50 ml .5 x TBE, .5 g agarose, with 2.5 ml ethidium bromide. Ethidium bromide is a fluorescent dye that intercalates between the bases of DNA. When the gel is put in a UV light box and exposed to UV light, the ethidium bromide bound to the DNA will fluoresce and report where our DNA bands are. Each band represents a population of DNA of a specific size. The size of each digested fragment is then determined by comparing how much the bands traveled compared to the DNA of known sizes in the control standards.

Rf = distance band traveled / total distance of dye front

The leftmost lane is the ladder control, and the four lanes on the right are comparing different samples of DNA treated with restriction enzymes. Each band represents a DNA fragment of a specific size.

Flow Cytometry In A Nutshell

2011-08-22T12:22:00.000-07:00

FACS was used in the trial treatments of leukemia last Saturday's post.

Fluorescence-activated cell sorting analysis, or FACS, uses a flow cytometer to separate individual cells in a heterogenous suspension based on epitope type. Fluorochrome-labeled antibodies are added to the cell sample. The antibodies bind to specific epitopes on or within the cell. The fluidics system delivers a stream of cells or particles one at a time through an interrogation point, where it passes through a laser. A cell traveling through the laser beam scatters the beam’s light forwards and sideways as a function of its size and granularity, respectively. When the laser beam strikes cells labeled with fluorescent-labeled antibodies, the fluorescent dye becomes excited and fluoresces at a unique wavelength. The intensity of the scattered and fluorescent light is collected and filtered by the optics system, recorded by the detector which translates the light into a quantifiable electrical impulse that can be represented graphically as a dot plot or a histogram.

We can use different fluorochromes at the same time, as long as their emission peaks are far enough apart for us to easily distinguish. A peripheral computer can instantaneously analyze the forward and side scatter light and fluorescence to identify the characteristics of individual cells and separate them into different subpopulations by charging each droplet with either a negative or positive charge, depending on the intensity and wavelength of fluorescence, as they leave the stream. The droplet is deflected either to the right or left by charged electrodes into one of three sample tubes. Intensity and wavelength of fluorescence and be plotted in a two dimensional box plot, where subpopulations in the sample can be distinguished by looking at two parameters.

2D results with two different parameters makes visualizing cell populations so easy!

A histogram measuring the frequency of celled labeled with antibody A is plotted on the y-axis of the two-parameter box plot, and another histogram measuring the frequency of cells labeled with antibody B is plotted on the x-axis. The box plot in this example shows two subpopulations, distinguished by the intensity of fluorescence f protein expression.

New Cure for Leukemia?

2011-08-21T17:30:00.000-07:00

Leukemia is the cancer of white blood cells.

The Penn scientists targeted chroniclymphocytic leukemia (CLL) by hacking a harmless version of the HIV virus to hack T cells in order to kill cancer cells. In previous studies, the cancer-killing cells died out quickly after infusion, but in this study, the genetically engineered cells multiplied a thousand-fold and were sustained for over 4 months.

Let's go over the study first. Three patients with chemotherapy resistant tumors had their blood drawn, separated, modified, and cultured. These patients underwent lymphodepleting chemotherapy, and their blood was reinjected. Endpoint assays were conducted a month after reinjection.

They did the same thing in mice, and the cells of interest were sustained for over six months, although I'm not sure whether the same monthly cycle was repeated. It doesn't say. But the cells of interest reached levels of up to 95% of white blood cells, up from 2.3-4.46% (figure 2). After an initial decay with first-order kinetics, the CART19 cell numbers stabilized between three to six months after reinjection. The fact that the cell levels were sustained after four months is at least some evidence the body can remanufacture the CART19 cells on their own.

What is most remarkable, however, is that the cells of interest seem to be able to remanufacture themselves within the body. In the third patient, flow cytometry showed that there were CAR19-expressing T cells with an absence of B cells 169 days after infusion. This is remarkable, since, "previous studies have not demonstrated robust expansion, prolonged persistence, or functional expression of CARs on T cells after infusion."

Figure 2 from the study showing levels of CART18 cells after infusion. Click to enlarge.

"There were no significant toxicities observed during the 4 days after the infusion in any patient other than transient febrile reactions. However, all patients subsequently developed significant clinical and laboratory toxicities between days 7 and 21 after the first infusion...With the exception of B cell aplasia, these toxicities were short-term and reversible. Of the three patients treated to date, there are two complete responses and one partial response lasting greater than 8 months after CART19 infusion according to standard criteria." The only side effect these three patients suffered was fever. One was hospitalized for a week, and another went into remission for 10 months.

In fact, "one of the preclinical rationales for developing CAR+ T cells with 4-1BB signaling domains was a projected reduced propensity to trigger IL-2 and tumor necrosis factor–α (TNF-α) secretion compared to CAR+ T cells with CD28 signaling domains (7); indeed, elevated amounts of soluble IL-2 and TNF-α were not detected in the serum of the patients." The cells infused into the patients were designed specifically to avoid a cytokine storm and to circumvent the donor's immune system.

"In our preclinical studies, we found that large tumors could be ablated and that the infusion of 2.2 × 107 CAR T cells could eradicate tumors composed of 1 × 10^9 cells, for an in vivo effector-to-target (E/T) ratio of 1:42 in humanized mice (8), although these calculations did not take into account the expansion of T cells after injection." In mice studies, billion cell tumors were ablated.

The three human patients had trillion cell tumors weighing around 1 kg before the infusion of CART19 cells. They all showed great progress, with the third patient surpassing others by 40:1. "Using the estimate of initial total tumor burden (1.3 × 1012 CLL cells) and the observation that no CLL cells were detectable after treatment, we achieved a marked 1:93,000 E/T ratio. By similar calculations, an effective E/T ratio in vivo of 1:2200 and 1:1000 was calculated for UPN 01 and 02 (table S6). Therefore, a contribution of serial killing by CART19 cells combined with in vivo CART19 expansion of >1000-fold likely contributed to the powerful antileukemic effects mediated by CART19 cells."

The trials for the three patients were financed by Alliance for Cancer Gene Therapy.

Source: Chimeric Antigen Receptor–Modified T Cells in Chronic Lymphoid Leukemia

(the study) and MSNBC.

No Science Sunday: GMSoccerPicks

2011-08-21T14:00:00.000-07:00


Best Soccer News and Write Ups. Ever.

GMSoccerPicks is a must-read blog for you soccer fans out there. He's got some real insight as an avid soccer fan, and his commentaries should not be missed. The blog is updated daily, so if you're into soccer, bookmark it!

You can also join the hundreds of people following @gmsoccerpicks on Twitter and get soccer news as it happens, or like GMSoccerPicks on Facebook to get soccer write-ups in your newsfeed.

Enumerating Bacteria In Lab

2011-08-19T11:46:00.000-07:00

Serial dilutions allow us to do viable cell counts or total cell counts.

Serial dilution and plating can determine the amount of viable cells in a culture. Serial dilutions allow a discrete number of colonies of bacteria to grow, whereas concentrated cultures may contain billions of bacteria per milliliter. In serial dilutions, smaller dilutions are repeated in succession, and the dilutions can be multiplied to obtain the total dilution. Thus, serial dilutions are more practical than doing the total dilution in a single time. For example, if I have a 100 ml bacterial culture, I can add 1 ml of it to 99 ml of water, add 1 ml of the first dilution to 99 ml of water, and then add 1 ml of the second dilution to another 99 ml of water. I end up with a 10-2 X 10-2 X 10-2 = 10-6 dilution of the original bacteria culture. I can then plate .1 ml of the final dilution on growth medium. The goal is to dilute the culture so that, when plated, the number of bacterial colonies is discrete and each colony arises from one viable bacterial cell. We can use the number of viable cells in the undiluted culture by dividing apparent colony-forming units with the product of milliliters used and the dilution factor. For example, if 150 colony-forming units were counted on the plate that was streaked with .1 ml of the 10-6 dilution, there is about 150 / (.1 ml X 10-6) = 1.5 x 109 bacteria/ml in the original culture. This method is useful because I am using only a small portion of the original culture, and large volumes of solution are not required for many-fold dilutions.

The Petroff-Hausser Counting Chamber can also be used to validate cell counts. The cell suspension is vortexed and a drop is applied to the chamber with a Pasteur pipette. Etched squares on the surface of the chamber representing specific areas and volumes are then examined under high magnification. Count the number of bacterial cells per chamber cell and multiply to obtain the concentration of cells per milliliter.

The turbidimetric method indirectly determines the quantity of insoluble particles in a liquid by comparing light transmittance in reference to a standard solution. A spectrophotometer shines a specific wavelength of light at the sample. Insoluble particles suspended in the sample will absorb and the incidental light, decreasing the amount of light transmitted to the photocell. Optical density is the measure of the turbidity of a solution, and it increases as the concentration and size of the particles increase. For example, as the concentration of bacteria reaches about 107 cells per ml, the liquid medium will appear cloudy or turbid.

Aλ= log10(Io/I) = εbc

The absorption of light is described by the Beer-Lambert Law, where A is absorbance, Io is the intensity of light incident on the sample, and I is the intensity of light transmitted through the sample. Beer’s Law states the optical density is proportional to the concentration of the compound in the solution, c, and the light’s path length, b. Thus, the concentration of bacteria in a pure culture can be determined if the molar absorbtivity, ε, and the path length, c, are known. OD600 refers to the optical density of a sample when the incident light has a wavelength of 600 nanometers.

Evolution and G6PD Deficiency

2011-08-18T21:20:00.000-07:00

Evolution takes place over thousands of years, when I asked about evolution, I was looking for an answer that had the same scope. I wasn't not talking about the past few decades, but the past few thousand years. So while some made very valid comments on botox and the nebulous cultural standards of beauty, the answer I liked best looked at the bigger picture and had some specific examples to support his thoughts.

M Fawlful made a good point about many genetic diseases becoming apparent later in life. These inherited diseases have no effect on the mating fitness of an individual. M Fawlful also stated that individuals heterozygous for the disease can actually be more fit and produce more affected progeny versus unaffected homozygotes. It's why some populations in Africa have sickle cell anemia; G6PD deficiency confers resistance to falciparum malaria (one of the biggest infectious killers in Africa).

The thing is, in developed countries, our environment is no longer selecting for any physical trait in particular. It is no longer putting a selective pressure against the unfit, because humankind has changed the environment to fit its needs.

We have made homes with air conditioning, built supermarkets and awesome hospitals, developed vaccines against polio and tetanus, for example. These allow everyone to live and thrive, regardless of their physical fitness or their potential skills as a hunter/provider.

A lot of you mentioned ugliness or perceived physical beauty was irrelevant when we're talking about sexual fitness, but I'm still not convinced. An individual's preference, influenced by upbringing or whatever, doesn't have as much influence on the evolutionary progress of a species. People are instinctively drawn to people who look a certain way. Having a symmetrical face is a sign of physical fitness. Having wide hips and large breasts is a good indication of a female's fertility, for example. That's what I mean by good-looks, but I digress.

Another good point made by many of you is that mutations are always popping up in our genomes, and these mutations lead to birth defects and weird traits like a long neck or whatever. Given that these mutations are the substrate on which evolution can take its course and new ones constantly pop up in every generation, ugliness and new genetic diseases can never be completely eradicated.

Ultimately, M Fawlful, I picked your comment out of the many great comments by Bellingham, Ghevrix, GMSoccerPicks, Inverse, Procras, Gareth Thomas, DS, Bersercules, Mekkor, Clueless Dolphin, Equalz, Electric Addict, Lars, Twist of Events, H., Maxe's Maze, Natural One, ason31, neversettleforsecond, Michael Westside, Randall A., Shaw, Timothy Bowen, Kid Shuffle, convictus, and last but not least, Bulletproof Zombie. To everyone who contributed to this very interesting discussion, thank you. To the people who have followed this website since July, thank you for your continued support. TheTruthAboutGenetics.com has 200 followers now.

Sources linked directly above.

Question of the Day

2011-08-17T09:25:00.000-07:00

Why hasn't evolution gotten rid of the ugly and genetically dysfunctional individuals? It's survival of the fittest, right? For the past hundred thousands of years, wouldn't ugliness and genetic diseases be slowly weeded out?

M Fawlful won the 8 GB SD card. Congratulations!

Huntington's Disease Explained Simply

2011-08-15T14:05:00.000-07:00

Cells in our body (except sperm cells and eggs) have two copies of every gene, one copy from your father, and the other from your mother. Genes are like blueprints or instruction manuals that tell the cell how to make proteins, the building blocks of the cell. Thus, genes and the proteins they encode for determine everything about the cell: how it grows, what it looks like, how it will respond to signals from its environment. Changes, or mutations, to these genes will cause changes to the proteins and affect the cell, much like a word-change in a sentence will change its meaning.

In Huntington’s Disease, a repetition of a CAG sequence in the gene encoding for the protein Huntingtin makes it clump together in our brain cells, ultimately making the brain cell die. For each CAG sequence in the genetic blueprint, the cell incorporates, one after another, an extra glutamate, a building block of protein, into Huntingtin. Longer repeats of the CAG sequence mean more glutamates are incorporated into the protein. It’s like a blueprint of a house that normally instructs an architect to build a chimney on the roof. One chimney is fine, but if the blueprint has an error and tells the architect to build 40 chimneys on the roof, the house would likely collapse, ruining not just the house, but damaging the area around it. Houses built with 50 or more chimneys would be even more unstable and cause more damage. Chimneys, and glutamate, aren’t inherently harmful, it’s their improper incorporation into houses and cells, respectively. In brain cells, the more glutamates in Huntingtin, the more protein clumps form, more severe the damage, and the lower the age of onset. This explains the variable age of onset of the disease, or the age at which symptoms arise; different people have different amounts of the CAG repeat.

The mechanism of the disease is still being researched, but here’s what we do know. The repetitive glutamates in the Huntington protein change the shape of the brain cells, affecting their function. The glutamate sends signals that constantly over-excite brain cells. Their overexcitement leads to cell damage, and ultimately cell death. Changes in the breakdown of nutrients will lead to the production of toxic chemicals known as free radicals. The regions of the brain that regulate movement, impulsivity, and learning are most affected in Huntington’s Disease. As a result of brain cell damage and death, Huntington’s have trouble controlling their movement, with rigid joints, difficulty chewing and swallowing, involuntary tics and writhing movements called chorea. Cognitive manifestations include impulsiveness, lack of empathy, memory loss, ultimately leading to dementia. These symptoms become progressively worse as time goes on.

The disease is dominantly inherited. Only one bad copy of the gene from either the mother or father will result in Huntington’s Disease. Children of people affected with the disease have a 50% chance of getting it from an affected parent, irrespective of whether the other parent has a normal copy of the gene. If both parents have Huntington’s Disease, offspring have a 75% change of being affected by the disease.

Source: Annu. Rev. Neurosci. 2007. 30:575-621

No Science Sunday: Wine Edition

2011-08-14T15:00:00.000-07:00

It's Sunday, the day of rest! I like to relax on Sundays and read books with big bag of Flaming Hot Cheetos and a few glasses of good wine.

Got a date or going to a fancy dinner? Looking to spend 9 or 10 bucks on the best bottle of wine to enjoy with your company? Go for a wine made in Portugal as a general rule of thumb. Given Portugal has the lowest wages in EU, they're pump out good wines at a comparatively low price.

25 bucks? No thanks.

Yesterday, I was at a wine shop looking for a good bottle of wine for a professor of mine, and I overheard a lady smugly complaining about how she is so sensitive to sulfites in "lesser wines" and it gives her headaches. I could tell she had no idea what she was talking about. Sulfite allergies manifest in asthmatic or anaphylactic reactions, never headaches. I guess some people like throwing around buzzwords they hear once at a wine tasting at a bar or read about Consumer Reports. Anyways, the salesman offered her a Conundrum, which she happily got.

Conundrum is definitely expensive, and I am not sure if they're worth the money, given that there any many superior wines priced a little less. I got an Evolution from Oregon for myself. It's as good for only 15 dollars. I certainly can't tell the difference. YMMV.

Unrelated note: saved 70 bucks today by changing my own engine and cabin air filters myself.

Regulation of Morphology of Corn Smut, Ustilago maydis

2011-08-14T09:03:00.000-07:00

Basidiomycota, in contrast to other fungi such as Ascomycota, produce basidia that yield four sexual spores called basidiospores. U. maydis is part of this phylum. Its mating-type is determined by a tetrapolar system with two unrelated loci, a and b. There are two idiomorphs for the a locus, a1 and a2. Haploid U. maydis cells have either the 4.5kb a1 locus with genes mfa1, pra1, and rfa2, or the 8kb a2 locus with genes mfa2, pra2, lga2, and rga2. mfa1 and mfa2 encode pheromone precursors, pra1 and pra2 genes encode pheromone receptors for the a2 and a1 pheromone, respectively. rfa2, lga2, and rga2 are thought to function within mitochondria. The pheromone encoded by one idiomorph will bind to the receptor of the opposite cell type, activating a signaling cascade that induces G2 arrest and the formation of conjugation hyphae. The b locus contains two genes, bE and bW, and regulates the switch to the pathogenic filamentous stage, as well as tumor induction and the formation of teliospores. The complex mating-type regulation is not specific to U. maydis, and other Basidiomycetes such as Schizophyllum commune and Coprinus cinereus.

Promycelium undergoes meiosis to produce saprophytic haploid cells. When in contact with corn, these sporidia exchange pheromones and become conjugative hyphae. These fuse to form a dikaryote, which is able to being intracellular invasion. Tumors are induced in which the fungi proliferates. Spores are formed and spread in the air and form a promycelium.

Higher fungi, like U. maydis, make ideal genetic models because they are easy to mate, transform, and select for. Observing metabolism, virulence, genotype is easier because they tend to be linked to readily apparent morphology. U. maydis’ relatedness to animal cells makes their study even more relevant to humans. Not only do does it have microtubule organization, nuclear migration, and nuclear envelop breakdown like in humans, U. maydis has homologues of Homo sapiens proteins that other, “higher” genetic models lack, such as Brh2, a BRCA2 (Breast Cancer Type 2 susceptibility protein) homologue. In vivo studies of Brh2 made it possible for geneticists to understand the function of BRCA2 in DNA repair and tumor suppression in humans. There is no doubt of U. maydis’ importance as a genetic model to study other complex mammalian cell processes.

No pictures. No sources. Only excellence.

Ethics

2011-08-13T00:14:00.000-07:00

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Yesterday, I asked whether you would kill an innocent girl to cure the world of HIV/AIDS, ultimately saving millions of lives. Given that HIV and AIDS kills 6,500 people every day, leaving millions of children as orphans in Africa alone, is the killing of one innocent person justified?

I couldn't kill one person to save millions of lives because doing so means I have to ask myself, "How far would I go? How many people would I kill to save millions?" Let's ask the question again, except this time, you have to kill ten innocent people to cure the world of HIV/AIDS. Would you still do it? What about killing a hundred? A thousand? Many of you justified killing one person to save millions, but would you kill thousands of people? At what point would you stop and say, "Alright, this isn't ethical anymore."

To the people who said they would kill the girl yesterday, how many people would you kill to cure the world of AIDS?