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  • Number of Reads in 454 Library

    What's the average/range of the total number of reads in 454 library?

  • #2
    The number will depend on the type of chemistry, the number of regions you are using (e.g., whole plate, quads, etc.), loss of quality that you willing to tolerate within the sequencing, the quality of your starting material, etc.

    Just to throw out some numbers our last titanium run with 4 different experiments (samples) on it gave us:

    136538 reads at 553 bases with std. deviation of 332 bases.
    392301 reads at 298 std. dev. 137 bases
    369737 reads at 318 std. dev. 154 bases
    391092 reads at 325 std. dev. 154 bases

    So roughly 1.3 million reads of around 300 bases or ~400 M base pairs.

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    • #3
      I really depends a lot on what you're doing. Our first run with a bacteria on the titanium got 1,371,195 reads, avg 397.4, std. dev. 135. In total 505Mb.
      But now we're doing runs with array enrichment and we get about 100Mb less per run. Although I've been told that a new protocol for array enrichment will perform much better.

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      • #4
        Which enrichment system are you using? I was thinking of using Nimblegen, but it seams too expensive, so I am starting with some test with the Combimatrix array first. Anyone using Combimatrix for seq enrichemnt??

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        • #5
          We're using the Nimblegen enrichment. We're doing the enrichment in-house so that makes it a lot cheaper.

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          • #6
            I was thinking of doing the same thing, but my concern is if it is possible to tag the samples before doing the enrichment. My target is <<5Mb, so I was thinking of using the Nimblegen array with a high tilling (many copies of the same few probes targeting 100kb per sample) so that I could enrich many samples all together.
            I was wondering if you ever tried anything like that?

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