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Has anyone experienced failed 454 FLX+ runs when using MIDs and flow pattern B? We have had 3 runs fail (control beads run fine) when using MID adaptors. These are rapid shotgun libraries. I have repeated library construction using regular adaptors and they run well.
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In what way have the runs failed? -
The first 2 runs(MID 31 and MID 32)
had no "key sequences". I remade the libraries with different MIDs (MID 13 and MID 23) and they seemed to work--although there are some anomolies in the actual assembled sequence--there is an 8 base sequence complementary to the MID sequence that appears thoughout and was tagged by the assembler as a possible primer.
The 3rd library key-passed at 96%, but gave only 50K very short (175bases) reads in 1/4 plate (MID 34). I remade this library without MID and it ran well (240K reads of avg 770bp)
We haven't assembled this one yet.Comment
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