Rover Exiting Crater To Continue Martian Marathon

Riding with Robots writes "The robotic geologist Opportunity has nearly reached the rim of Victoria Crater, which it is leaving after a year of exploration inside. Rover handlers decided to abandon attempts to approach the crater's cliff walls when they saw a power spike similar to the one that preceded a broken wheel on its twin, Spirit. Opportunity is already making do with a stuck robotic arm. The mission's manager said, 'Both rovers show signs of aging, but they are both still capable of exciting exploration and scientific discovery.' Opportunity is set to continue trekking across the Meridiani Plains of Mars, even though its wheels have already seen 10 times the use they were designed for. Meanwhile, Spirit has survived yet another harsh Martian winter to produce another striking panorama." Adam Korbitz notes other Mars-related news that funding has been approved for the Search for Extra-Terrestrial Genomes (SETG) Project. The project was one of 15 selected to receive funds through a NASA research opportunity program. The stated goal of the proposal is to "develop a PCR detector for in situ analysis on other planets, most immediately, Mars. This instrument is so sensitive it should allow the detection very low levels of microbial life on Mars, and will determine its phylogenetic position by analysis of the DNA sequence of the genes detected in situ."

From the f'ing article

[Anonymous Coward]

16S RNA gene PCR, the most sensitive detector for life on Earth

This detector is an amplification strategy called the Polymerase Chain Reaction (PCR) that is based on artificial replication of DNA. PCR is a technique which is used to amplify the number of copies of a specific region of DNA, in order to produce enough DNA to be further analyzed. In order to use PCR, one must know the exact DNA sequences which lie on either side of a given region of interest in DNA. One need not know the DNA sequence in-between. A DNA sequence is the precise order of appearance of 4 different deoxyribonucleotides. The 4 components are: Adenine, Thymidine, Cytosine and Guanine, abbreviated A, T, C and G, respectively. The arrangement of this 4-letter alphabet is the DNA sequence.

The PCR strategy for life detection emerged from the exploration of the diversity of life, which revealed about 500 Ã'universal genesÃ" that are carried in the DNA of every known living thing on Earth (7). The gene that has changed the least over the past 3-4 billion years is the 16S (or the related eukaryotic 18S) ribosomal RNA gene. Ribosomal RNAs are the main structural and catalytic components of the ribosome, a molecular machine that translates RNA into proteins (8,9).

It is the slow rate of change of the 16S gene that makes it the best detector of life. Within the ~1500 nucleotides of the 16S gene, there are multiple 15 to 20 nucleotide segments that are exactly the same in all known organisms (8). These regions of the 16S gene are essential for its catalytic activity and have remained unchanged over billions of years (8).

  The technology of PCR involves adding stable 15-20 nucleotide long DNA primers, a stable enzyme nucleotide triphosphate monomers, and a simple heat pump that thermally cycles 20-30 times in 2 hours. To amplify 16S genes from a crude sample, universal DNA primers from the ribosomal RNA gene that are about 18 bases long, oriented towards each other, and about 1000 bases apart are added to crudely purified DNA isolated from an environmental sample (for example, 1 ml of sea water or 1 gram of earth). For the ribosomal genes, the DNA primer 5Ã GTGCCAGCAGCCGCGGTAA 3Ã which corresponds to nucleotides 515 to 533 of a ribosomal gene, and 3Ã TTCAGCATTGTTCCAWYGGCAT 5' which corresponds to the base pairing complement of nucleotides 1492 to 1510 are added to an extract prepared from soil (M, Y, and W are codes for mixtures of two such nucleotides necessary to capture all 16S genes). Upon heating to 95ÂC and then cooling to 55ÂC, these DNA primers pair with their complement on each DNA strand, even if there are only a few DNA molecules in a sample. After heating to 75ÂC, the DNA polymerase will polymerize the nucleotide monomer components also in the tube to duplicate the DNA strands. There will now be four strands, where originally there were only two. If one repeats the thermal cycle with all the same components in the same tube, now there will be eight strands; repeat again - now 16, etc. Thirty cycles will produce one billion (230) copies of the original sequences. Because the DNA polymerase enzyme used derives from a thermophilic microbe, it can survive repeated cycles of heating to 95ÂC. The amplified DNAs from the PCR can be analysed for size or DNA sequence. PCR will even amplify complex mixtures of 16S ribosomal RNA genes from communities of organisms in environmental samples. Thus, PCR with DNA primers corresponding to the conserved elements can be used to amplify DNA from any species more than a billion fold, without need to isolate, culture, or grow the organism in any way (9).

Re:PCR? With what primers?

[Eicos]

They would use random primers. A DNA hexamer (six-base sequence) is sufficiently long to serve as a PCR primer, but short enough that it would take only 4096 different types of molecule to comprise all possible sequences. Of course, we don't want the sample DNA to be plastered in our primers, so we'll pare those 4096 down to a handful, at least one of which, in any sample sequence of significant length, will nonetheless find somewhere to anneal. Once we've gone through enough cycles, it's likely that we'll have amplified at least some segment of the sample DNA. Then, getting the reaction contents purified and sequenced is simply a matter of applied microfluidics.

Re:PCR? With what primers?

[SlashBugs]

Use random primers, just like you do for reverse transcription when you want to pick up all the RNA sequences in your sample. The reaction's efficiency would take a hit, but if all they want to do is detect DNA (or maybe even sequence a few very short sections) it could probably be made to work.

A bigger problem is the enzyme used in the PCR. IANABiochemist, but I'd expect the PCR to only work if the Martian bugs hava genomes based on double-stranded DNA chemically very similar to ours.

There are plenty of stable nucleotides that could work as components of DNA but, for some reason, aren't used in Earth's life. Ditto chirality: Using the same constituent atoms, one can build almost identical but left- or right-"handed" versions of molecules. For some reason -- probably just chance -- Earth's life is based on "lefthanded" molecules, meaning that we can't produce or consume right-handed molecules. For example, if we synthesise right-handed sugars (easy for a chemist to do, but expensive), they have the same chemical composition, melting point etc, but the structure is such that our enzymes can't use it as a source of energy. Heck, even the sequence of any DNA scooped into the chamber will be important, as if influences the reaction conditions you need for the PCR to work.

If there is life on Mars, this test would only be able to detect it if Martian life is spookily similar to our own. Which would, I'll admit, be even more exciting than just "life on Mars" because it would hint toward evidence of Panspermia or possibly some sort of fundamental rules about what life is able to look like.

Re:PCR? With what primers?

[drerwk]

Kind of like the microscope on Phoenix? http://www.nasa.gov/mission_pages/phoenix/news/phoenix-20080814.html [nasa.gov]
100nm resolution. DNA however is only 3nm wide.

Re:PCR? With what primers?

[Dr.Dubious DDQ]

Their working hypothesis (spelled out on the linked page) is that early in the development of microbial life there is (they claim) a statistically relevant chance that microbes developed on either Earth or Mars could have survived transfer from one planet to another via "meteoric exchange", which there would appear to have been a lot more of back ~3.5billion years ago when the first signs of modern-style microbes appear in the geological record.

Their assumption is that regardless of whether microbial life originated on Earth and possibly got blown to Mars during a major meteor impact, or vice-versa, if there are microbes growing in both environments now they'll be related.

I was going to say that you don't necessarily need specific bases for DNA amplification - there are some "whole genome amplification" techniques now that use a mix of small "random" primers to get amplification of (hopefully) most or all of the DNA in a sample rather than just one gene.

However, the description of the project does explicitly say they're planning to try to amplify 16s ribosomal DNA sequences, which are very handy for phylogenetic analysis of known terrestrial prokaryotes:

"a set of DNA oligonucleotides that also universally detect ribosomal genes (906-922F=GAAACTTAAAKGAATTG and 1407-1391R= GACGGGCGGTGWGTRCA, where K = G or T, W = A or T, and R=G or A) but prime within the 519 to 1492 region amplifed in the first step will be used. "

I'm a bit skeptical of the "universality" of "universal" primers, especially as to their usefulness after ~3,000,000,000 years of divergence. On the other hand, unlike some of the previous tests a positive result from this experiment would be very unambiguous if they can rule out contamination.

Re:PCR? With what primers?

[waferbuster]

yep, based on the single datapoint of earth life using DNA, it's reasonable to expect and look for ET DNA. However, after rolling a die once and getting 4 it would be similarly reasonable to expect subsequent rolls to also be 4.
Extrapolating based on a single datapoint is shaky at best. But without alternative substances to check for, DNA seems reasonable.

This reminds me of the old saying about when holding a hammer, everything looks like a nail.

Re:More Mars color BS

[Teilo]

Here is a before and after, if anyone cares:

http://tinyurl.com/5mon9r [tinyurl.com]

Re:More Mars color BS

[sighted]

Here's a page that attempts to rebut this: http://www.donaldedavis.com/2008%20new/CLRMARS.html [donaldedavis.com].