Kun:LabNotes/ExonomeSeq/2008-1-9
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Exp.1 Sequencing library construction for the E55kv1/GM10835 amplicon 450bp band[edit]
Just realized that I should do Mme I digestion prior to other enzymatic reactions.
Mme I digestion of amplicons[edit]
dNTP ampilcon dUTP amplicon dUTP USER digestion DNA 20ul 10ul 15ul NEBuffer 4 3ul 3ul 3ul SAM(1mM) 1.5ul 1.5ul 1.5ul Mme I 2ul 2ul 2ul H2O 3.5ul 13.5ul 8.5ul
37C 1h -> 80C 20min -> 4C hold
File:2008-01-09-Lib-construction.png
Digestion was not complete. I added 2ul more enzyme, 37C 2h -> 80C 20min.
PAGE purification of the 250bp bands.