User:Meng Xiao He/Notebook/fall08/2008/11/02

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Ligations/transformations

 * XmaI heat killed
 * vectors treated with ant. phosphatase
 * phosphatase killed


 * fragments column purified


 * used NEB T4 ligase b/c quick ligases have PEG

DH5a, Kan+Xgal:
 * TOPO + each flank's original PCR
 * pUC19 + each flank
 * pUC19 neg control
 * 4ul per transformation

pir+ electrocomp cells, LB spread w/ Gm+Xgal:
 * each pER21 prep + each flank
 * pER21 neg control

Rx

 * 8ul DNA mix (2:6), 1ul ligase, 1ul buffer
 * 20 min ligation


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