Alan+Y

=Week Four Wiki! 6/3-6/8: Alternatives to PCR =

===The highlight of this week's classes was probably the PCR lab. PCR represents a major innovation that has changed biological research and advanced our understanding of organisms' genomes. No longer do we have to use the "shotgun method" of transformation via prokaryotes. We now have a small machine that can quickly replicate millions of copies of a target sequence. I am still amazed that humans have developed something as ingenious as PCR. Although PCR doesn't always work, at least we now have a method that can ideally give scientists a bunch of copies of a known sequence. This has greatly facilitated research.===

===In recent years, scientists are exploring ways to copy DNA segments other than PCR. This is really important to. We always want our methods and technology to progress as our knowledge becomes deeper and deeper. We must not let laboratory inadequacies hinder our exploration. Though PCR has bestowed numerous benefits on researchers, scientists have been working on alternatives to PCR. These alternatives are by no means perfect, but it is interesting to take a look at what scientists have been developing.===

===Q-beta is a bacteriophage, and this virus naturally contains an enzyme replicase that can copy RNA-->RNA. In other words, Q-beta is an RNA-->RNA virus. In the same sense that RNA Polymerase II in humans attaches at the promotor/TATA box, replicase attaches at a sequence of nucleotides called MDV-1, which can be extracted from the bacteria species //Plasmodium facoparum//.===

===Scientists can use Q-beta when they want to make millions of copies of an RNA sequences that is complementary to the DNA sequence under study. This is sort of analogous to the concept of cDNA, which we learned in class. The only difference is that in Q-beta Relicase Amplification, the RNA serves as a template for //another// RNA strand.===

===Thus, scientists can construct primers that consist of the MDV-1 recognition sequence and the first few nucleotides of a particular RNA strand. They can mix these primers with the enzyme replicase and RNA extracted from cells. The result is millions of copies of an RNA sequence.===

===Q-beta Replicase Amplification still has some glitches. However, scientists are working to overcome these glitches because of some of the benefits of Q-beta Replicase Amplification. These are: (1) Q-beta can amplify //RNA// (2) Q-beta can operate at a constant temperature (37 C) and (3) Q-beta products can be stained with ethidium bromide, and the intensity of the stains correlate with the logarithm of the amount of RNA. In other words, if a product stains 3 times as intensely as another product, there is roughly 1000 times more RNA in the first product.===

===Unlike PCR, LCR is highly specific (the nucleotides don't keep extending) and is used primarily to detect SNPs, single nucleotide polymorphisms. For this reason, LCR is used a lot in the detection of diseases (like sickle cell anemia and certain STIs) in which there is a single nucleotide that is abnormal. Like PCR, LCR uses three different temperatures for each round of replication.===

=== 1. Synthesis short DNA probe sequences. Unlike PCR, you need 4 of them for LCR, 2 for each strand. LCR is really specific: only the nucleotides between the probes will be replicated. The probes should be synthesized on either side of a potential SNP. ===

=== 4. Ligation: This part is different. Since the probes are adjacent to each other, ligase is used to join them--but only if the annealing was done right. In other words, if there is a SNP, then the probes won't line up and ligase won't be able to join the adjacent probes. Thus, LCR can only amplify sequences that are exactly like the two end probes on each strand. PCR, on the other hand, can replicate any sequence after the primers. (Note: sometimes DNA polymerase is also used in this step to replicate the nucleotides between the probes if the probes are not synthesized to be adjacent to each other.) ===

A more detailed description of LCR (like hard core) can be found here: [].
=== What's interesting about LCR is that it is already a very viable option for detecting SNPs. For instance, in one study, LCR fared better than PCR in diagnosing patients with Chlamydia (an STI). Although the study said that the difference between LCR and PCR was not statistically significant, the P-value was 0.125, which is moderately small even though it is greater than the 0.05 cutoff. Thus, LCR is a very powerful tool in diagnosis. ===

Link of the study: []
=Week Three Wiki! 5/27-5/31: Nanotechnology= === When we go to Robbin's Park to study the streams, we are taking a first step into the study of ecology, how living systems interact with each other and their environments. I feel like as the study of ecology gets more advanced, its scope broadens, and the units to be studied get bigger and bigger. We are now looking not just at population dynamics, but whole ecosystems, and even the entire biosphere. It's interesting that in other subdivisions of biology, the focus of study gets smaller and smaller as the field gets more advanced--subdivisions like virology and immunology. So small has our focus gotten in these fields that many researchers are now looking not just into microbiology, but into nanotechnology. ===

=== Nano is really small. Like 10^(-9) small. Way smaller than the macroinvertebrates we looked at in class under microscopes, and sometimes even smaller than red blood cells. One of the most important applications of nanotechnology is in medicine, where such tools can be used to help fight diseases. Some of these applications are listed below. ===

1. Cancer
===Nanotechnology can furnish us with minuscule "robots" that can float around in our body (like our interstitial fluid) and identify and latch onto cancerous cells through specific binding probes. Then, it can inject a poison, rip open the cell membrane, or use some other method to kill that cell. The biggest challenge with cancer therapy via nanotechnology is constructing probes that are specific to the binding sites of cancerous cells (and not normal cells) at the molecular level. One way to accomplish this might be to take advantage of the computer processing capabilities that can be installed within a nano-robot. This computer can help figure out or detect whether a cell is cancerous or normal.===

Here is a patient testimony of nanotechnology treatment of cancer from Lankenau Health Center--an institution that's fairly close to home, in the Main Line area.
media type="youtube" key="bvwqxmrmBCo" width="560" height="315"

2. Providing Oxygen
===Tissue damage or any other conditions that hurt red blood cells can deplete one's oxygen supplies. Here, nanotechnology can be used to boost oxygen levels. A proposed structure for such a nano-robot I found is this:===

=== "a sphere with an internal diameter of 0.1 microns (100 nanometers) filled with high pressure oxygen at ~1,000 atmospheres (about 10^8 pascals). The oxygen would be allowed to trickle out from the sphere at a constant rate (without feedback). Diamond has a Young's modulus of about 10^12 pascals [so it can endure the stress]" ([]). ===

===From the description above, it's apparent that a lot of chemistry and physics are involved. The article above also mentions that the ideal gas law, PV=nRT, would probably need to be used to calculate pressures at certain temperatures and volumes in order to maintain a constant rate of oxygen transfer. As for physics, it suggests that the most preferable material to use to contain the oxygen gas at ~1000 atmospheres is probably pure diamond because of its high Young's modulus.===

===The most complex issue with this application of nanotechnology is that we need to find a safe, allergy-free material to encapsulate the whole robot. We don't want patients to undergo allergic reactions or side-effects after the robot is introduced.===

Other pictures:


=== In class throughout the whole year, Mrs. Lil repeatedly used sickle cell anemia as something that touches upon many biological concepts. Well nanotechnology might be a solution to sickle cell anemia, as it can help raise oxygen levels in the blood. Of course, such a nano-robot might not be able to do much about blood clotting, but at least it's a start! ===

3. DNA Repair
===In cells where the DNA has been damaged (which sometimes leads to oncogene stimulation and cancer), nanotechnology again provides a possible solution. The nano-robots can help by going into the nucleus and physically/chemically repairing the base pair damages. Or, these robots can also serve to check the DNA for errors. You might be thinking--don't we have enzymes to do this? Well, nanotechnology can act as a double check, and it can also handle situations where either the errors are so great that the enzymes cannot cope or if the DNA-related enzymes are dysfunctional.===

4. Fighting diseases
=== Using similar methods as mentioned above, nanotechnology can be used to fight pathogens, both bacteria and viruses. These nano-robots can be employed to combat and kill these pathogens. Thus, nanotechnology may be a possible alternative to anti-biotics, providing a solution to the "superbug" problem. === === One of the motivations for my post this week was Henry's post last week about anti-biotic resistant bacteria. I think that nanotechnology might be a possible solution to the anti-biotic question, and I have high hopes for the future of nanotechnology. ===

=Week Two Wiki! 5/20-5/24: Integrated Pest Management=

===If I had to make a list of things I could live without, stinkbugs and mosquito bites would definitely make that list. Though they certainly are a nuisance to ordinary people, pests, especially those of the phylum Arthropoda, are particularly annoying to gardeners and farmers. They can ravage hard-earned crops, infest nearby habitats, and disrupt native ecosystems. However, as mean as the pests may seem to humans, we cannot reciprocate with heavy-handed tactics. We must control, NOT necessarily eradicate, these pests in a way that is cost-efficient and least disruptive to the local ecosystems.===

2. Identify the pest (VERY crucial!!). This determines what methods will be appropriate.
=== 3. Decide whether the pest has reached levels that are unacceptable. Remember, it's really hard to ERADICATE pests, so take action only when pests have reached unacceptable numbers, and not as soon as you come across one. ===

__Preventative Practices__
=== It's just as important to prevent pest infestation as it is to control it later on. Ways to prevent the arrival of pests include things like (a) growing strains that are innately resistant to a certain pest and (b) not introducing foreign species. ===

Using mechanical barriers is the most desirable of all the control methods because it does the least harm to the ecosystem (and it is often cost-efficient!). These include:

 * ===Straight-up hand-picking===
 * ===Traps===
 * ===Barriers and fences===
 * ===Setting up anything that the pest is averse to (like maybe shaded areas)===

These methods must be used with caution. These include:

 * ===Introducing natural predators.===
 * ===Introducing natural substances/chemicals that ward the pests away.===
 * ===Introducing fungi or bacteria that are disruptive to the life cycles and reproductive cycles of the pests.===

Of course, when worst comes to worst, you'll be forced to use pesticides with care.
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= = === The purpose of IPM is to remind everyone that pesticides are not the only way (and certainly not the safest way) to bring pests under control. Pesticides can be dangerous (think DDT), and should be used as a last resort. ===

=== Strict, explicit, and specific definitions and guidelines of IPM have not been established yet. This is partly because appropriate IPM methods naturally would vary from region to region, and the pest levels deemed "unacceptable" also vary. ===

Please comment, I look forward to seeing what you think!
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=Week One Wiki! 5/13-5/18: The Gaia Hypothesis=

====__**Introduction:**__ I used to wonder if global climate change, and specifically global warming, is reversible. If humans make the planet hotter and hotter on average, does the earth have natural mechanisms to counteract these changes? Can the earth bring its own temperature back down to normal by somehow invoking colder weather?====

Does the earth operate via homeostatic feedback loops? Can the earth as a single entity somehow respond to external changes almost like a living organism?


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Fun fact: Gaia is the Greek goddess representing "mother earth" and she gave birth to the Titans and the Greek gods.


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=History of the Gaia hypothesis:=

==== Hardcore version: The Gaia hypothesis is an ecological hypothesis proposing that the biosphere and the physical components of the Earth (atmosphere, cryosphere, hydrosphere and lithosphere) are closely integrated to form a complex interacting system that maintains the climatic and biogeochemical conditions on Earth in a preferred homeostasis. ([]). ====

Here is a video of Lovelock explaining his hypothesis:
media type="youtube" key="44yiTg7cOVI" width="420" height="315" align="center"

==== I was VERY intrigued by this idea that the Earth is living. We are always told that rocks and wind and such are abiotic factors, and they are. However, if you take all the abiotic factors in the world in sync with all the biotic factors, perhaps you will get a larger system that mimics life, and yet supercedes all life forms yet. ====

1. Oxygen and carbon dioxide levels have a set point, a preferred homeostasis.
====2. Global temperatures have set point. I do, however, want to qualify this statement. I personally don't believe that the Gaia hypothesis outright contradicts the global warming trend. It just states that the earth has mechanisms to oppose change. However, if humans constantly spew out harmful toxins into the environment, the earth will inevitably get unhealthier.====

=__**Initial criticisms of the Gaia hypothesis:**__= ====There were many criticism of the Gaia hypothesis in the 1970s (surprise, surprise--a new idea meeting skepticism). Most of these were very technical; however, one major complaint is very relevant to our AP Bio course this year:====

====Well, does it? There's still debate. However, scientists initially argued that basically, if the earth is living, then it abides by natural selection. If the environment itself is being naturally selected for, then the organisms inside those environments are going to be undergoing some pretty messy natural selection. Sure, changes in the environment that are "bad" will be selected against--but then what about the organisms in those environments?====

====James Lovelock has suggested in more recent times that the earth indeed does undergo a form of natural selection IN CONJUNCTION with the organisms, in that **ORGANISMS THAT HELP BRING THE EARTH BACK TO ITS HOMEOSTATIC SET POINTS WILL INCREASE THEIR CHANCES FOR SURVIVAL AND BE SELECTED FOR**. In other words, organisms that can help the earth are actually benefiting themselves. It's kind of like __**coevolution**__.====

5. Uses energy -- I suppose as a whole, the earth can transform and utilize energy to do work. The earth may not be using energy consciously, but when light heats up the oceans--that's energy!
====6. Grows and develops -- Arguably...the earth certainly has changed since the Big Bang. For instance, the oxygen revolution we learned about in class was when photosynthetic bacteria started churning out oxygen.====

8. Respond to the environment -- YEAH!
=The Gaia hypothesis today:= ====In 2006, there was a conference about the Gaia hypothesis. By now, the Gaia hypothesis has earned much more credit than in the 1970s, and James Lovelock was actually awarded for his contributions. Here is the website that was made for the conference:====