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Old 02-18-2014, 06:12 PM   #1
jwag
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Default Anyone had luck with Fosmid library prep kits?

Hi -- I was wondering if anyone has had any luck with making 40kb jumping libraries using a Fosmid kit. I was looking at this NxSeq 40kb mate pair cloning kit . But really I have no idea what a good kit would be-- the whole process seems like there's a lot of room for mistakes. It will be for Illumina sequencing. Thanks!
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Old 02-20-2014, 08:01 PM   #2
krobison
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What are you sequencing? You might be better off using long reads (PacBio or Moleculo), which won't be 40Kb but given the recent successes (such as assembling an entire Drosophila chromosome arm)

This is the only kit like it on the market, and it is indeed an involved protocol -- you are first making a cosmid library and then converting it to a sequencing library.
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Old 02-21-2014, 09:14 AM   #3
jwag
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I'm using Illumina technology for sequencing. I looked into PacBio, but the genome I'm trying to sequence is 1.6 GB, far too expensive for PacBio currently even only at 1X coverage. I might just try to find a company that does Fosmid libraries...
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Old 02-21-2014, 09:43 AM   #4
GenoMax
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Do you anticipate having difficulties (repeats, ploidy considerations etc)? Is there a related genome available?

Depending on how "finished" you want the genome to be, you may be able to get a lot done with normal libraries.
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Old 02-21-2014, 09:47 AM   #5
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The genome of interest is highly repetitive (estimated 21% tandem repeats, 42% repetitive). Unfortunately, there isn't a related genome available. I wanted to be able to do structural rearrangement analysis and comparative genomics between different species, which I would think requires a high degree of genome completion.
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Old 02-21-2014, 09:59 AM   #6
GenoMax
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Well .. in light of that your choice is going to be limited. Hopefully you can at least do a few SMRTcells on PacBio or other nanopore based technologies would become more generally available this year.

I know that oomycete genomes have been done (and they were certainly problematic back in the day).
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Old 02-21-2014, 10:02 AM   #7
jwag
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I might just wait until PacBio or SMRTcells become more affordable... as of right now it's about $6k to do 1X coverage of PacBio for a 1.6 GB genome.
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Old 02-23-2014, 06:59 AM   #8
Genohub
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Hi jwag,

Doing a project search on Genohub for exactly 1x coverage on a 1.6 Gb genome yields the following PacBio service providers:

https://genohub.com/shop-by-next-gen...fb2229a23f817b

Genomax -- we're hoping too that nanopore tech comes online this year. By chance, are you one of the lucky few that has early access to the MinIon ?

- Genohub
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Old 02-24-2014, 07:10 AM   #9
GenoMax
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Quote:
Originally Posted by Genohub View Post
Genomax -- we're hoping too that nanopore tech comes online this year. By chance, are you one of the lucky few that has early access to the MinIon ?
I do not work in the lab but I know at least one person locally who was turned down. So not everyone who applied probably made it.

Perhaps we should do an anonymous poll and see how many on SeqAnswers got selected?
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