User:Jeffrey Kim/Notebook/Halogenated natural products from uncultured bacteria/2008/12/12

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454 sequencing

  • Prep appears to be ok although we ran into problems with concentration in pools 1 and 4. A retry of the initial library construction indicates that the DNA is of high quality and in sufficient amount to proceed. Sequencing preparation continued according to mfg. protocol without problems. Final titration of emulsion PCR concentrations also looked fine. Final sequencing produced a high proportion of mixed reads. I believe that this is due to overloading of the plate based on a 10% bead recovery after counting. We will retry the experiment with fewer beads and with a lower concentration of DNA during emulsion PCR.