Re: [CCL] Sepia bandensis

[Patrik D'haeseleer]

Here's an excellent article on keeping and breeding Sepia bandensis:

http://www.advancedaquarist.com/2005/9/aafeature

A single cuttlefish seems to require at least a 20+gal reef tank. Definitely significantly more challenging than raising some zebrafish, but not out of reach for a dedicated amateur with experience with reef tanks.

Patrik

On Tue, May 2, 2017 at 4:30 PM, Shanee Stopnitzky <shanee.s...@gmail.com> wrote:

I'd be interested in all of it but husbandry in particular, I used to work on cuttlefish vision and kept a bunch of the little darlings. I haven't worked with this species but cuttlefish in general are non-trivially difficult to keep healthy in closed systems! 25th works for me. 

Best,

Shanee

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<><    <><                 )     <*III><  

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<><        <><  <><     ) ) 

'the fact remains that political frontiers are impervious to our verbal cultures, while the substantially nonverbal civilization of playfulness crosses them with the happy freedom of the wind and the clouds.' ~ primo levi 

On May 2, 2017, at 3:48 PM, Craig Rouskey <craigr...@gmail.com> wrote:

Hello Folks,

I wanted to put out some feelers to see if anyone was interested in having a meeting to discuss the Genome/Transcriptome experiments for dwarf cuttlefish. I was thinking we could have a meeting Thursday May 25th at 7pm...

Does that work for folks?

Anyone interested in Cuttlefish Husbandry (Wranglin'); RNA-Seq/DNA-seq; RNA editing, bioinformatics, etc. could join.

Let me know what you think...

Craig

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[Maria Chavez]

I'm interested.  Where to host the meet up?

Sent from my iPhone

On May 2, 2017, at 3:48 PM, Craig Rouskey <craigr...@gmail.com> wrote:

Hello Folks,

I wanted to put out some feelers to see if anyone was interested in having a meeting to discuss the Genome/Transcriptome experiments for dwarf cuttlefish. I was thinking we could have a meeting Thursday May 25th at 7pm...

Does that work for folks?

Anyone interested in Cuttlefish Husbandry (Wranglin'); RNA-Seq/DNA-seq; RNA editing, bioinformatics, etc. could join.

Let me know what you think...

Craig

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[Patrik D'haeseleer]

Having been involved in many genome sequencing projects, keep in mind that de novo sequencing, genome assembly, annotation and analysis of a higher eukaryote is still a VERY ambitious project, typically involving a consortium of multiple academic labs.

It's definitely feasible, but this would be a very significant effort, that should not be undertaken lightly.

I would recommend we get our feet wet by sequencing and annotating some novel bacterial genomes first. There is *plenty* of work to be done on that, and that's at least an order of magnitude easier than a large eukaryote.

Patrik

On Wed, May 3, 2017 at 10:15 AM, Craig Rouskey <craigr...@gmail.com> wrote:

Ken,

I think that the majority of the aquarium work will happen at BioCurious. Cephalopods are known for their capacity for RNA editing, and a recent article looked at the K+ ion channel diversity in Sepia officianalis. Basically, they raised their cuttlefish at different temperatures, dissected out the brains, and noted diversity in the K+ channel transcripts. Then (the fun part!) they expressed the variants and did functional assays on them.

I'd like to run the whole transcriptome under different environmental conditions (x3 variables, maybe looking at salinity, or pH) see which proteins are most edited, pick a few, express and perform some f(x)al studies...but that comes later.

Also, Sepia bandensis does not have a reference genomic DNA sequence. We can sequence the genome as well for reference, or compose our RNA transcriptome de novo...

DNA and RNA-seq will contribute a lot to the scientific community, but it's a lot of data and we'll need a lot of hands contributing to the annotation of the genome. Some of it should be pretty straight forward, the rest, not so much. Our experiment will result in a ton of data that will need to be sorted through. Coding/Non-coding RNA structures, variable transcripts, etc.

I've put out a call to some friends at Amaryllis nucleics to help with the tissue processing. Basically, for $200 a sample, we send frozen tissue and they do the processing. The processed RNA/cDNA will be sent off for sequencing through Illumina (I'm thinking a 30M x 176bp read depth, but I have mroe research to do into that). The majority of the working is the husbandry and dissecting of our cuttles. Also, Amaryllis will help us with RNA-seq, not ICE-seq which is a method to detect Inosine, pre-"G" transition. We may want to price out ICE-seq.

Here are some papers...

Craig

On Wed, May 3, 2017 at 9:30 AM, Ken Litchfield <litchfi...@gmail.com> wrote:

OK, so why cephalopods as the phylogenetic group to work on?

Will we be having a marine aquarium in CCL?

Other compatible organisms to round out the ecosystem? Can the luminous dino's handle a cephalopod aquarium?

Thanks Ken

On Wednesday, May 3, 2017, Craig Rouskey <craigr...@gmail.com> wrote:

Exactly, Kevin!

Sent from my iPhone

On May 2, 2017, at 11:43 PM, Kevin Brennan <kb...@stanford.edu> wrote:

I’m guessing it’s a practical model organism for cephalopods?

Kevin 

From: Craig Rouskey <craigr...@gmail.com>
Date: Tuesday, May 2, 2017 at 10:59 PM
To: Cheryl Ching <cheryl...@gmail.com>
Cc: Rachel Linzer <rachel...@gmail.com>, Maria Chavez <ishy...@gmail.com>, Jaddua Ross <jaddu...@gmail.com>, Ccl Igem <ccl-...@googlegroups.com>, cclabs <counterculturelabs@googlegroups.com>, biocurious <biocu...@googlegroups.com>, kbrennan <kb...@stanford.edu>
Subject: Re: [RVC] Sepia bandensis

It's the transcriptome and we're studying the patterns in its RNA editing capabilities.

Sent from my iPhone

On May 2, 2017, at 10:09 PM, Cheryl Ching <cheryl...@gmail.com> wrote:

Am I missing something here? What is the significance of the dwarf cuttlefish/dwarf cuttlefish genome?

On Tue, May 2, 2017 at 9:50 PM, Craig Rouskey <craigr...@gmail.com> wrote:

I'll set the meet up for both CCL and BioC... we can do a zoom scenario...

C

Sent from my iPhone

On May 2, 2017, at 9:28 PM, Rachel Linzer <rachel...@gmail.com> wrote:

I'm also interested!

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[Patrik D'haeseleer]

On Wed, May 3, 2017 at 10:54 AM, Craig Rouskey <craigr...@gmail.com> wrote:

I was hoping to do a crowd funding campaign...

I'm currently pricing out the costs for all of the work.

My usual recommendation for projects such as this is to just ignore sequencing costs!

Next-Gen sequencing has gotten so cheap that the actual sequencing cost winds up being only a tiny fraction of the total cost of the project. Of course, that mostly holds for academic projects where you have to price out the hundreds to thousands of person-hours of effort required to do all the sample processing and downstream analysis as well.

But I think the same principle works for a DIY effort as well: if you are able to cover the cost and effort involved in housing the squid and processing all the samples, and you are able to form a large enough coalition of DIYbio experts who are willing to dedicate a not-insignificant chunk of their lives to this project... then raising a few thousand dollars to do the actual sequencing will be minor in comparison.

Patrik

[Patrik D'haeseleer]

On Wed, May 3, 2017 at 12:01 PM, Craig Rouskey <craigr...@gmail.com> wrote:

Nope.

On May 3, 2017, at 11:12 AM, Cheryl Ching <cheryl...@gmail.com> wrote:

Is there a way to do this without killing them?

The genome sequencing at least could be done on a small tissue sample without necessarily killing the animal.

However the RNA editing that makes these animals so interesting seems to be predominantly found in the central nervous system. So if you really want to study this aspect, I don't think there would be any other option than by killing an animal.

At a very minimum, I think we would need to set up an Institutional Animal Care and Use Committee (IACUC) to do any work like this at CCL - including academic and outside community representatives. I think squid are strictly speaking not legally covered under the Animal Welfare Act (since they are cold-blooded invertebrates). But they are highly intelligent creatures, and we do have a moral obligation to minimize their pain and suffering.

We also have an obligation to our own members - and to some extent our surrounding community - not to work on projects that some people may find morally abhorrent. I'm not sure how well we could reconcile this project with our supporters for the Vegan Cheese project, for example...

Patrik

[Maria Chavez]

Hey folks I just posted a meetup for this on the 25th at BioCurious
and CCL. There will be an online Zoom meetup for anyone who wants to
attend remotely. This should be alot of fun to dive into!

Maria

On Wed, May 3, 2017 at 2:46 PM, Craig Rouskey <craigr...@gmail.com> wrote:
> Just spoke to the head of IACUC at Novartis, and she said she'd volunteer
> time to review an IACUC protocol for S. bandensis.
> She also said that liquid nitrogen is the probably the best way. Her name is
> Laura Tandeske.
>
> Craig
>
> On Wed, May 3, 2017 at 2:33 PM, Craig Rouskey <craigr...@gmail.com>
> wrote:
>>
>> I think you're right, John. Liquid nitrogen euthanasia is probably the
>> least painful and does the most to preserve our sample.
>>
>> Sent from my iPhone

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