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you

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[Music]

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thank you for coming everyone just a

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brief update on the title as you may

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have read so I changed the title to

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evidence for Institute iron cycling from

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focusing just on the let's autotrophs

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because I think the iron cycle in

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general makes for a more interesting

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story in my system so to get everyone

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oriented chocolate pots is in

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Yellowstone and it's in the northwest

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corner right along the given river

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denoted by the blue X and here we have a

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couple pictures of hot springs we have

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the oops there we go so we have the vent

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pool up here the flow path and then

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chocolate pots is very interesting hot

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spring because it's circum neutral pH

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mildly thermophilic and contains a high

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amount of iron and silica in the spring

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water from a biological perspective were

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interested environments like this

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because of the new and recent

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discoveries of certain neutral pH like

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environments on Mars and the evidence of

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relic hot spring-like features and if

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we're able to nail down evidence for

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internally cycling iron base metabolisms

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in environments like chocolate pots then

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this could in theory be applied to some

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of these relic features on Mars so we

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have two main hypotheses going forward

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with this study the first is that there

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are native active iron reducing

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microorganisms present in the chocolate

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pots event and also that there are lipid

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atrophic iron oxidizers which are also

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actively cycling iron and in addition as

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let the autotrophs they are fixing

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carbon debt yeah fixing co2 and able to

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supplement other sources of carbon for

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the heterotrophic community

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so backing up a little bit some of our

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initial work with materials from

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chocolate pots we ran a series of iron

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reducing enrichment culture studies and

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this is a graph showing the repeated

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transfer of these cultures for over four

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years and they've been able to sustain a

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fairly high level of iron reducing

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activity and through both 16s and meta

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genomic sequence data we've identified

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sequences that are related to known iron

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reducing taxa including a Geo vector and

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mealy factor moving one step towards

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choosing apart the infant we ran another

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series of short duration incubations

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using just the native sediments and

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amended with and without additional

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electron donor and the things I'd like

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to point out here is in this feature

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which they're the that figure which

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corresponds to material collected from

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the hot spring vent you have an almost

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equivalent level of iron reduction

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activity regardless of electron donor

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source whereas if you move a few meters

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away at site three there's still a

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measurable amount of activity of the

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organisms using their native carbon

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source but it's greatly reduced so this

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led us to the question of what is this

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internal carbon source is it coming from

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chemoautotrophs is it from the

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cyanobacterial community is it breakdown

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of plant or animal to trade detritus

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from the external environment whereas

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the carbon coming from so and we have

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16s data from this as well and several

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Oh to use related to an organism

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Ignasi bacterium while it is not

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implicated as an iron reducer it is very

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similar to this newly or a factor which

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I identified in the enrichment cultures

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taking a look now at the in Scituate

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vironment we collected a series of core

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samples moving down the flow path and

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the 16s data from these samples revealed

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sequences related to known with

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autotrophic iron oxidizers sitter

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oxidants and again this relative of a

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potential iron reducer Ignasi bacterial

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ease so magic and ohmic work flow I'll

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cover in brief you start out with a very

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complex community like chop the pots get

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all the DNA sequence it put it back

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together separate it out into bins which

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are you can think of as being different

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taxa within the community and then what

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we're really interested in is what is

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the metabolic potential of some of these

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these bins or taxa and we're taking a

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targeted meta genomic approach looking

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at which genes are appear to be involved

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in iron cycling and which ones are

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involved in carbon fixation so here we

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have the community of two metagenomes

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that I assembled from chocolate pots one

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is from water filtered from the vent

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pool and another one is a combined

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assembly from three of those sediment

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cores that I mentioned and once again

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you see bins related to this known with

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the autotroph and this possible iron

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reducer so thank you to Dave Emerson for

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the lovely introduction - I see - I will

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not be covering that in my talk because

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I found no homologues that in my system

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but I will be talking about this other

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electron or extracellular electron

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transfer system a pouring wrapped

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version where you have a pour in in

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opening in the outer membrane of the

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cell and a cytochrome which fits into

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that and is able to transfer electrons

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into or out of the cell depending on

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whether it's an iron oxidizer or I

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reducer

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and there's two homologs in particular I

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want to point out there's this MTO a

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system which is identified in said er

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oxidants and there's this geo factor

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like pouran cytochrome system and it's

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also been identified in that Ignasi

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bacterium organism these are just

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several different search tools that you

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can use hidden Markov models blastp

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homology many different ways to identify

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these genes but one of the more

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important things that we're interested

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in is the genomic context of these genes

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that genomically you need to find a pore

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in encoded next to one or several of

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these cytochromes okay we're looking for

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multiple co2 fixation pathways but for

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the sake of brevity I'm only going to

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cover Rubisco today so to quote my

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advisor if you identify a bin with an ET

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system and Rubisco that's a pretty good

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smoking gun of an organism that may be

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involved in a litter litho autotrophic

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lifestyle and we very similar search

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parameters for these genes so here I

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have a summary of all of the metagenomic

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bins from these two assemblies in red we

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have all of the bins that contained some

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model EEG system in yellow we have all

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of the bins that contained a hit for

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Rubisco and what we're mostly interested

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in is these bins that contained both

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systems and may be involved in chemo

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little otter trophy and then down here

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at the bottom are bins that were not

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involved in any of it

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so bringing this back to the two

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organisms that I've been mentioning

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throughout this talk the sitter oxidants

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and the ignominy she really is

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I just wanted to summarize the results

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in terms of those two organisms so up in

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gray we have the published genomes of

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these organisms and against utter

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oxidants containing this MTO system and

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Rubisco and the two related

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Ignasi bacterium and me leroy vector

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containing only that geo vector like TCC

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system whereas looking at the two

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metagenomes

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here we have a sitter oxidants relative

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that all of a sudden has a PCC system

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and we have these Ignasi bacterias that

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have MgO and Rubisco which is really

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interesting that sitter oxidants which

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is a known with autotrophic iron

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oxidizer contains PCC which so far has

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only been implicated in iron reduction

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so perhaps this is a relative that is

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able to carry out both iron oxidation

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and iron reduction and for the Ignasi

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bacteria Li's as I mentioned before has

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an e ET system and Rubisco so perhaps it

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is capable of carrying out a little odd

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so to wrap things up once again we have

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a highly active dissimilatory iron

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reducing community at the chocolate pots

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event and it is able to sustain itself

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using only the native carbon source

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we have metagenomic bins that are

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related to known and potential iron

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cycling microorganisms we have the

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positive identification of homologs of

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genes involved in both these eeet

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pathways and carbon fixation pathways

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and as a side note we're in the process

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of identifying lipid biomarkers from

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these materials to try and nail down a

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possible source of the carbon that's

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present in statue and one last thing I

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would like to mention so I only covered

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very briefly the model systems that

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we've been looking at but we've also

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expanded our search to look for non

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model ET systems and we're also looking

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at all of the known carbon fixation

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pathways in these systems and that I'd

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like to thank the organizers for the

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conference my funding my committee

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members who have helped with this

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project Eric and Eric and my colleagues

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back at UW Madison particularly Xiaomei

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hay who without her meta genomic

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expertise we would not have been able to

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do any of this so with that I think I

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so we definitely have time for a couple

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of questions Tim yeah yeah I really

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enjoy the Yellowstone toxemia yeah

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interesting it's a cool area

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so what non-model systems do you have in

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mind exactly as far as the Ichigo yeah

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yeah so again a lot of this credit goes

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to Xiaomei and her computer programming

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skills but we have scripts that were

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able to search our meta genomic

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sequences for the Keene binding site

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motifs and any genes that have that

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appear to be multicam cytochromes we

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then look for genes proximal to that

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that are predicted to be an outer

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membrane pore in so the ones that aren't

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homologous to any of the models but

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still seem to fit that same type of

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system with a pouran and a cytochrome

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what about the EPI or GOP ly any

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considerations about yeah so that's

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something that I've considered and we've

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discussed as a lab quite a bit but

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something that I I still need to

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investigate with these systems yeah

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thanks yeah welcome thank you so we

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still have time for another question if