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you

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[Music]

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thank you very much Beth and Alexis for

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the opportunity to share my latest

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research in progress with you guys here

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today so today I'm going to talk about

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some meta genomic data from a tropical

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thank you

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so we've already heard about the process

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of serpentinization multiple times today

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so I won't take too much time here to

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belabor the point however as far as

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astrobiology is concerned it's

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essentially a process where we've got

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minerals and water coming together and

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producing life molecules hydrogen

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methane and small organic carbons and so

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from an astro biological point of view

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that has implications for both the

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origin of life here on earth and

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potentially life on other planets one of

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the best characterized sites of

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serpentinization is the lost city

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hydrothermal field which is located off

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of the mid-atlantic ridge and the

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communities there are dominated by

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methane cycling archaea on the interior

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of the chimneys and they're called

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velocity Misano sarsen alleys and then

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the exterior of the vent chimneys are

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slightly more diverse and contain

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aerobic methane atrophic bacteria and

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what this suggests is that methane

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cycling by microbes is incredibly

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important to communities in velocity

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this is a map of some sites that our

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research group either has projects at or

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collaborations with other groups at and

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we've been studying serpentinization in

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a continental setting for years and so

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these results from the lost city beg the

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question is methane cycling by microbes

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important in these continental sites of

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certain ization as well so methanogens

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have been detected both at Oman and

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delivery and ophiolite

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in Italy using either 16s or meta

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genomics data sets in Portugal the table

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Enzo filet in Canada

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and the coaster angel feel light in

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California they have not been detected

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and that's either from 16s datasets

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metagenomic data sets are a combination

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of the two and I just want to use this

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moment to plug a poster that's at this

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evening if you'd like to hear more about

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the table m sophia light please see

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poster 33 11 by a technician in the

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Brazelton lab and so for the remainder

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of the talk I would like to talk about

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this one site in Costa Rica and whether

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or not we see methane cycling going on

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there so the Santa Elena alveoli is

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located in the North Pacific corner of

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Costa Rica a paper by Sanchez Rio at all

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looked at various geological and

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geochemical and microbiological

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parameters at a variety of sites this

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location today I'll be talking about

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sites located right here so two sites in

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particular we've got spring 9 and a very

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close proximity

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upstream site for a background sample

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spring 9 has the typical travertine

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calcium carbonate deposits that you'd

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expect when the high calcium fluids from

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serpentinization come into contact with

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the inorganic carbon in the air and

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despite they're very close proximity of

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a couple of meters from each other they

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have different geochemistry

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a spring 9 has a pH of 11.5 and an

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abundance of hydrogen and methane

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whereas the background sample still has

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an elevated pH but it's much lower and

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has less of these volatiles and bio

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energetic calculations have suggested

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that Mehsana trophy and Mehsana genesis

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are both favorable at both of these

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sites so now to get into a little bit of

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the microbiological data a lot of

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continental serpentinization sites have

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either no or very low abundance of

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archaeal toxa and so here we looked at

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some quantitative PCR data and i've

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highlighted here the two samples that

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we'll be talking about in this talk and

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well the archaea

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well archaea are less of

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and the bacteria they still make up a

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substantial portion of the microbial

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communities which is contrary to what

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we've seen in other sites to explore the

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methane cycling a little bit further I

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mind some 16s rRNA amplicon datasets to

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look for potential methane cyclers here

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we're looking at the bacterial

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communities and I did not want to do

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that I just want to orient you here that

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the units are the percent of the total

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bacterial community but the axis goes

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from zero to ten percent and that's

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because these were not in very high

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abundance at all we see some aerobic

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Masana Tropes in spring nine and a

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slightly higher diversity and higher

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abundance in the background sample but

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it still makes up less than 1% of the

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total community the archaea tell a

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slightly different story again we're

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looking at the percent of the total of

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peer community but here the the access

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ranges from zero to 100% in the warm

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colors were looking at methanogens and

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in the cooler colors their putative

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mechanic tropes and so here we can see

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that in our background our in our high

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pH spring 9 the methane cycling archaea

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make up between forty to eighty percent

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of the total archaeal community and in

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the background sample they're still

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significant making up about thirty to

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forty percent of that community so since

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not everybody in the room right now is a

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molecular biologist I thought I'd take

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just a few moments to explain some of

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the methodology for the data I'm about

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to present so we use meta genomic

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sequencing and we filter the natural

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microbial communities we then extract

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DNA and chop that up into little pieces

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that can be sequenced on a

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high-throughput sequencer the ultimate

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goal for metagenomics is to try to

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reassemble complete genomes of

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individuals from the environment and so

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we take these small little pieces of DNA

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and try to put them back together in

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larger chunks through assembly methods

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those larger trucks chunks of DNA are

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called con tags

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binning is the step where you try to put

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back together these genomes either

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complete or more likely near complete

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and so here this represents potential

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individual organisms where you can then

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annotate and predict their different

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metabolic pathways and so this is an

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example of the assembly from spring 9

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the high ph spring from this assembly we

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have eleven metagenomic bins ten of them

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were classified as bacteria one was

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classified as unknown which means that

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it wasn't even able to be determined at

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the domain level and the coverage

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represents how complete I'm sorry the

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completeness represents how complete

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these genomes are since we're trying to

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obtain ideally full genomes and so you

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can see here we have a wide range from

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17 to 83% completeness which of the

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lower end is is not great and so would

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these bacteria I should point out that

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number one none of these were identified

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as explicit Masana tropes and also we

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don't see any are keel bone here however

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as I mentioned before from this

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metagenomic data we can also not only

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look at who's there but what their

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metabolic potential is and so this is

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the Masana genesis pathway these numbers

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here all represent potential genes in

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the pathway the genes highlighted in

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blue were found in both the background

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and the high ph spring whereas the genes

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highlighted in orange were found

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exclusively in the high ph sample and i

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should note that there were no no genes

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involved in methanogenesis that were

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found exclusively in the background

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sample and so what we can see here is

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that we have complete meth an agenda

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this fast way in the high pH Springs

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also there were no I searched for

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bacterial Nathaniel genes in these meta

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genomes and none were found so as I

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mentioned there's been a lot of research

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going on in the past few years into the

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microbial diversity

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metabolism in continental

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serpentinization sites and from that

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there some models have been developed

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some things that are found almost

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universally is an abundance of these

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beta Proteobacteria and also anaerobic

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clostridia and this data set was no

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different there were an abundance of

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both of those groups one question that

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still remains is what the role and

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abundance of these archaea methanogens

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are in this system we still don't have

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an answer to that as they're found in

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some sites and not others but we're

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beginning to dive into that now so just

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to summarize the evidence for Monsanto

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Genesis at the santolina ophiolite

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include bioenergetics

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our keel 16s data both from qpcr and

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amplicons and Fulmer Masana Genesis

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pathway in the spring nine meta-genome

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however know our keel bins were found in

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that meta-genome and i just want to take

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a quick moment to explain that

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assembling metagenomes is much like

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taking hundreds of puzzles and spreading

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out all the pieces and trying to

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reassemble those puzzles again and so

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well in this go-around the puzzles

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didn't come back together very well

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there are other methods to try to try to

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improve those bins so my next steps are

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going to be to try to optimize the meta

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genomic assemblies and try to get more

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complete and representative bins

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additionally we have meta transcriptomic

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data for both of these samples and meta

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transcriptomes can tell you not only

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what genes are present in the

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environment like the metagenomes but

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they can tell you which ones are

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actively being expressed by the microbes

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so stay tuned with that I would like to

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thank my co-authors and collaborators my

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funding source of the deep carbon

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Observatory as well as everyone who

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helped out with data analysis sequencing

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Thank You Katrina we have plenty of time

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for questions if folks want to come up

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to the microphone or stand up and speak

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really loudly yeah that's very nice so

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one of the persistent issues in genome

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analysis is unassigned operating frames

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or open air nutrients with unknown

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functions do you find in your in your

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various samples from different

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environments greater or lower abundances

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of the hard to assign stuff that might

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that might indicate that whatever

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they're doing this environmental

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condition needs to make use of them yeah

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that's always a problem so anytime that

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you're trying to assign whether it be

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taxonomy or in this case function you're

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limited by the databases that exist and

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the databases are limited by what is

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already known and so I think even in

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e.coli about 40% of the genes are still

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hypothetical meaning Sara Dean but we

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don't know what it does and that number

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just goes up as you get into unexplored

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environment off the top of my head I

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don't know what percentage of our Dean's

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are hypothetical but it's definitely an

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issue cultured representatives can help

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us address that but is it is it fairly

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consistent across the different samples

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sources or philo types that's a great

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question I don't know the answer to off

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Catrina I have a question the fact that

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you see some of these archaea I wonder

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if it's related to other parts of the

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chemistry of these waters in addition to

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just the methane and hydrogen

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particularly what is the sulfate

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concentration like in these fluids yeah

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I know that some measurements were taken

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I don't recall what they were off the

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top of my head but we are beginning to

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look into some of these other electron

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acceptors at these sites to answer some

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of the questions any other questions for

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Katrina there we go I got one hey

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Katrina Thanks so do I understand it

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correctly that the methane cycling genes

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were found just in the random

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meta-genome but they did they weren't

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recovered in any bin no actually

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so the annotations were done on the bins

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which raises a question of the validity

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of some of the bins that those genes

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were all archeo but all of the bins were

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identified as bacteria and so I think

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what we likely have is some archaeal

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contamination within those bins so it's

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promising that we see the full pathway

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but I think the bins need to be

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reassembled to try to get up at a little