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DNA ligation: Difference between revisions
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#[[Tom Knight]] has read that ligase can inhibit transformation. By heat-inactivating the ligase, this inhibition can be avoided. However, according to the NEB FAQ, heat-inactivation of PEG (which is present in the ligation reaction) also inhibits transformation, therefore a spin-column purification is recommended prior to transformation if you are having problems. | #[[Tom Knight]] has read that ligase can inhibit transformation. By heat-inactivating the ligase, this inhibition can be avoided. However, according to the NEB FAQ, heat-inactivation of PEG (which is present in the ligation reaction) also inhibits transformation, therefore a spin-column purification is recommended prior to transformation if you are having problems. | ||
#Treating PCR products with proteinase K prior to restriction digest dramatically improves the efficiency of subsequent ligation reactions. <cite>Crowe-NAR-1991</cite> | #Treating PCR products with proteinase K prior to restriction digest dramatically improves the efficiency of subsequent ligation reactions. <cite>Crowe-NAR-1991</cite> | ||
#Using [http://probes.invitrogen.com/products/sybrsafe/ SYBR Safe DNA Gel Stain] is a safer, non-carcinogenic alternative than ethidium bromide. | |||
==Acknowledgments== | ==Acknowledgments== | ||
