1. An analysis of the microbial communities of the Mojave Desert serving as a terrestrial model for the environment of Mars Elaine P. Bryant NASA Spaceward Bound Mojave 2010

2. Why the desert? If microbes exist on Mars (or other planets) one factor they must cope with is an extremely dry environment – no liquid water.

3. Terrestrial deserts:

4. Zzyzx Mojave Desert Region Mojave Barstow

5. Overall research plan: Examine the microbial communities at each site by: collecting soil samples along a precipitation gradient analyzing the microbial community at each site comparing results to determine trends or patterns relating precipitation and the specified communities

6. March 2007 - 9 – Spaceward Bound Mojave Fieldwork: –Established 7 sample sites along precipitation gradient –Took 3 samples at each site Laboratory: –Performed two culture techniques –Prepared DNA for molecular techniques

7. Sample transect

8. Sampling sites: Site 15 Tehachapi Site 17 Cramer Junction Site 21 –

9. Sampling procedure: 1. Identify an appropriate area 2. Establish a 10m x10m sample area 3. Determine sample sites w/in sample area 4. Take sample from top 8 cm

10. Analysis at Zzyzx 1.Count plates – to determine the relative number of microbes per gram of soil as a function of precipitation 2.Biolog sole-carbon-source microplates – to observe differences in the utilization of carbon sources as a function of precipitation 3.Extraction of community DNA for molecular studies

11. Count plates  CFUs/gm soil SampleAnnualCFUs/g siteprecipitation200720082009 1525.04.4x10 6 1.5x10 8 1.5x10 7 1615.42.9x10 5 1.7x10 7 2.32x10 6 1712.02.5x10 5 1.5x10 7 3.16x10 7 1811.32.6x10 5 1.3x10 7 3.29x10 7 199.01.3x10 5 1.5x10 7 1.45x10 7 20~8.8NA1.3x10 7 2.51x10 6 218.62.7x10 4 3.0x10 6 2.68x10 6

12. Biolog sole-carbon-source micro plates

14. Molecular analysis Analysis based upon the microbe’s DNA: 1.extract DNA from the organisms in the soil. 2.make copies (amplify) a gene of interest - 16S ribosomal subunit, using Polymerase Chain Reaction (PCR) 3.Then either: a)Make clone libraries to associate specific clones with taxonomic groups  phylogenetic trees b)Denaturing Gradient Gel Electrophoresis (DGGE)

15. Cloning basics: Compare 16S rDNA fragments from all members of the microbial community –Make ‘lots’ of copies of a gene from one organism (cloning) –Identify (sequence) the nucleotide bases of the fragments –Compare sequence to existing known sequences –Determine community based on phyla –Compare communities from one site to another

16. Mojave data 2007 Bacteria

17. DGGE Pass DNA fragments through a gel containing compounds to separate the two strands of DNA (denature) – each resulting band represents a “taxon” or genus of organisms If two columns contain the same banding, those communities have the same taxons of organisms 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17

18. Summary of activities for SBM ’08: 1.Collect 3 soil samples/site 2.Inoculate 2 culture plates per site for count plates 3.Inoculate 2 Biolog s-c-s plates/site 4.Extract DNA samples

19. Research plan: Research plan: Sample soil along a precipitation transect in a desert Analyze microbial communities specifically Bacteria, Archaea & Cyanobacteria Culture techniquesMolecular techniques Count plates Compare results Biolog sole-carbon source micro plates Cloning the 1.5kb 16S rDNA fragment DGGE