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文献和实验TOP10 chemically competent cells
reduces transformation efficiency by about 3x the first time, and about 6x total after several freeze/thaw cycles. Measurement of competence Transform 50 μl of cells with 1 μl of standard pUC19 plasmid (Invitrogen) This is at 10 pg/μl
. Step 6: Centrifuge the cell suspension at 4,000 RPM in the GSA rotor for 10 minutes and resuspend the cell pellet in 1/50 volume of ice cold, sterile 85 mM CaCl2 in 15% glycerol w/v. Dispense in 100 mL aliquots and freeze cells at -80oC.
Preparing chemically competent cells
Materials Plate of cells to be made competent TSS buffer LB media Ice Glassware & Equipment Falcon tubes 500μl Eppendorf tubes, on ice 200ml conical flask 200μl pipetman or repeating
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