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==== Adaptor ligation reaction mix ==== * I prepare 10X diluted TruSeq adaptor (conc. after dilution = 1500nM) Index 1-10 by mixing 2ul of 15uM TruSeq adaptor with 18ul H2O. {| {{table}} border = 1 | align="center" style="width:200px;background:#f0f0f0;"|'''Components''' | align="center" style="width:80px;background:#f0f0f0;"|'''Volume''' | align="center" style="width:80px;background:#f0f0f0;"|'''half rxn''' | align="center" style="width:80px;background:#f0f0f0;"|'''11.2x rxn''' |- | End-repaired/A-tailed DNA||60.00||30.00||0.00 |- | Ligation buffer||30.00||15.00||168.00 |- | DNA ligase||10.00||5.00||56.00 |- | '''Total'''||'''100.00'''||'''50.00'''||'''20ul/rxn''' |- | Diluted adaptor (1.5uM)||10.00||5.00|| |- | '''Total'''||'''110.00'''||'''55.00'''|| |} :- Add 5ul of 1.5uM diluted Truseq adaptor to each tube ::- Final conc. of adaptor in 55ul ligation reaction is ~136nM :- Add 20ul of ligation reaction mix to each tube :- Mix well (spin down & pulse-vortexing 10X & spin down) :- 20C, 15min :- Continue to SPRI clean up immediately
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