Cavalcade of Mammals

What would be required to set up an inexpensive system for hobbyists to experiment with biology? Consider PCR for example. PCR requires heat stable polymerase, primers, nucleotides, buffer.

The DNA polymerase is easy to purify from E. coli carrying the plasmid. Grow bacteria containing plasmid expressing Taq DNA polymerase, boil, spin down denatured proteins, and you are left with the DNA polymerase.

Primers can be bought inexpensively–$0.35 per base, a pair of 18-mers cost less than the shipping. Though they are inexpensive only if one set gets used repeatedly.

The cost of buffer (NaCl, MgCl₂, Tris) is negligible.

Nucleotides are expensive up front, $150 for a set of dNTPs (dATP, dCTP, dGTP, dTTP), but this works out to about $0.06 per 50 ul PCR reaction.

Can nucleotides be prepared by a hobbyist? Nucleotides are easy to obtain-DNA is a major constituent of cells and is easy to purify. DNA + DNAase = dAMP, dCMP, dGMP, and dTMP. How can the trinucleotides be regenerated?

One route is to do it enzymatically using
polyphosphate:AMP phosphotransferase (PPT) and adenylate kinase (AdK) with polyphosphate (polyP) as the energy source (Resnick and Zehnder, 2000). It is not clear how the trinucleotide product would be separated and purified. Presumably different enzyme pairs could be used to regenerate the other dNTPs from the monophosphates.

These other enzymes could be cloned in E. coli expression vectors and purified either by tagging them with His₆ and using a Ni or Co resin. Or by cloning heat-stable isoforms from one of the extremophiles and using a one-step boiling purification like that used for Taq polymerase.

Update: Bochkov et al., 2006 describe a method of preparing dNTPs from digested DNA. DNA is digested with DNAase and Nuclease S₁. DNAase chews DNA into show oligonucleotides and the nuclease breaks them down to single dNMPs.

Then a crude extract of E. coli is prepared that contains the kinases to convert dNMPs to dNTPs along with the acetokinase. The kinases use ATP. ATP must be regenerated, and this is done using acetokinase with acetyl phosphate ($30/g) as an energy source. Combined dNMPs were converted to dNTPs with at least 86% regeneration and separated from reactants by chromatography on a Dowex 1×2 anion exchanger. The conversion was followed by thin-layer chromatography.

For PCR it may be possible to use

Last October, NASA’s LCROSS mission slammed a spent rocket booster then the LCROSS spacecraft itself into the moon. No debris plume was seen from Earth, but observations from LCROSS of the booster hitting indicate the presence of water on the moon. How much water? Most news accounts don’t say, but the Science magazine article does.

100 kilograms of water was detected from an impact that created a crater estimated to be 20 m wide and 3 meters deep. So 100 kg water in about 500 m³ of regolith = 0.1 g/kg. (Googled a reference giving 2.3 to 2.6 g/cm³ as lunar regolith density).

The article gives a higher estimate for water, 0.1% to 10%, higher than my crude 0.01% estimate. Which is great–enough water to extract easily and live off. Best news for space exploration in thirty years!

(Credit: NASA)

Interesting letter about the evolution of the glucocorticoid receptor from Joe Thornton, a biologist at the University of Oregon. Thornton’s lab has figured out some of the details on how this receptor evolved. Thornton’s writing this because creationist Behe is passing off a mangled version of his work as evidence for ‘intelligent design’.

The first Discover magazine blog post gives a good overview of what Thornton’s lab learned about the glucocorticoid receptor.

Here’s a web page at the The Jackson Labs site showing the most common mouse coat color mutations [link].

Here is white-bellied agouti with head blaze, greyish coat:

In a passing during a discussion of the lack of a system for naming viral strains and diseases, Greg Laden mentions that Ebola was named after the town in Zaire/Congo where it was first found.

That has to depress the local real estate market!

H1N1 flu is going around this year, an especially bad flu to which few people under 35 have much immunity. A bad flu year, but not in the same class as the 1918 flu.

The bird flu had potential to be very deadly, potentially as bad as the 1918 flu. The bird flu, an avian influenza A (H5N1) virus strain that emerged in Asia in 2003 had potential to turn into a very deadly human flu. A flu virus emerged that spread as an epidemic in birds or several species. Then it passed to humans–several times–and was very deadly, 60% fatal. While deadly, the virus that humans got was either wouldn’t transmit between people or did so only with great difficulty. Small clusters of people, usually of families that worked or lived with birds, came down with bird flu.

Bird flu needed only one or two mutations to become capable of causing a worldwide pandemic. It needed to pick up the property of easy contagion, human-human transmission. The first steps to contain bird flu in Asia failed. It spread around the world in wild and domestic bird populations and sometimes infected other animals. It has become endemic in bird populations this gives the virus many opportunities to infect humans and pick up mutations to allow it to spread in people.

So far we’ve been lucky, and the H5N1 avian influenza hasn’t turned into a fast spreading human flu epidemic. Perhaps this flu virus is a dead end of sorts and is incapable of jumping to humans. It does keep getting chances and infecting the occasional person so the CDC and WHO continue to monitor it. It has had many chances to become a human epidemic and so far hasn’t, and as time it becomes less likely to jump to humans. It looks like we got lucky.

We didn’t get so lucky with the capacitor plague…

Census data on median income change between 2000 and 2008 by way of USA Today:

US GDP in constant dollars per capita grew 9.7% over this period, so the country grew 10% richer and if it was distributed evenly everyone’s income would have grown roughly 10%. But not–instead most of the $$ went to a small slice of the population, and that doesn’t show up in this table.

I want a pet book worm!

The North Auckland worm, Spenceriella gigantea, reportedly glows in the dark brightly enough to read by!

In 2005, a Schweitzer et al. reported that they had found soft tissue in the marrow of a T. rex. bone. Examination of the tissue showed evidence of cell-like structures that could indicate preserved cells. Also stuff was squishy like protein. It if held up, an incredible discovery.

In 2007, Schweitzer et al. followed it up with more extensive analysis that indicated the presence of collagen I protein. Most exciting, they used mass spec to sequence a few fragments of dinosaur protein. It was quite similar to chicken collagen, and of course birds are the closest living relatives of dinosaurs.

Then in 2008, Kaye et al. analyzed a number of fossils and found similar traces, but interpreted them as bacterial colonies, as biofilm. They even saw the cell-like traces in a fossil ammonite, a creature without blood cells.

This year, Schweitzer et al. doubled down, publishing collagen I protein sequence from a hadrosaur.

In total the two papers found protein sequences that cover 169 aa of collagen protein sequence. Here are the overlapping dinosaur sequences with the gaps removed aligned to chicken sequence. The dinosaur sequences differ from the chicken sequence at five positions.

It will be interesting to see if these results hold up. I would like to see replication by a second lab. Most convincing would be two labs given the same unknown sample and both finding the same new collagen protein sequence.


>Tyrannosaurus rex
>Gallus gallus
>Brachylophosaurus
GATGAPGIAGAPGFPGARGAPGPQGPSGAPGPKGVQGPPGPQGP--------------------------
GATGAPGIAGAPGFPGARGPSGPQGPSGAPGPKGVQGPPGPQGPRGLTGPIGPPGPAGAPGDKGEAGPPG
GATGAPGIAGAPGFPGARGPSGPQGPSGAPGPKGVQGPPGPQGPRGLTGPIGPPGPAGAPGDKGEAGPSG
                   **                                               * 

--------RGSAGPPGATGFPGAAGRGVVGLPGQR-----------------------------------
PAGPTGARRGSAGPPGATGFPGAAGRGVVGLPGQRGETGPAGPAGPPGPAGARGSNGEPGSAGPPGPAGL
PPGPTGAR-GSAGPPGATGFPGAAGR---------GETGPAGPAGPPGPAGARGSNGEPGSAGPPGPAGL
 *

-GLPGESGAVGPAGPPGSR
RGLPGESGAVGPAGPIGSR
RGLPGESGAVGPAGPPGSR
               *  

Putting together the reading list for the seminar class this semester I ran across an odd corner of the human fertility industry that I never knew existed. I’m picking seminal papers in biotechnology and one of them is the 1992 paper, “Birth of a normal girl after in vitro fertilization and preimplantation diagnostic testing for cystic fibrosis” by Handyside et al. One of a pair of papers that first used PCR to test embryos for genetic disease so that the disease free ones can be implanted.

So I looked at where preimplantation diagnostic testing was available in Lexington. There is an IVF clinic that does preimplantation testing–not at the UK hospital, they seem shy of anything slightly controversial. What was odd, is ZDL, a company that sells fertility products for people to use at home, or send in for analysis. Home sperm counts, insemination devices, and more. Quite surprising!