New England Biolabs
Phusion High Fidelity DNA polymerase
M0530S
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I have used phusion high fidelity polymerase for amplification of DNA templates/genes needed for cloning. I got good quality amplification products which are error free. Quantity obtained was sufficient for cloning purpose. It is much better than Taq polymerase.
PCR- cloning of bacterial genes
50 ng of bacterial DNA
50 ng of total genomic DNA from P. aeruginosa was used to amplify target gene of interest in 50 microliter reaction according to manufacturer's instruction. Some of genes have high GC so I used 5X GC buffer in the reaction. Rest of the protocol and cycling conditions were as mentioned in the kit.
If you have high GC content, then use 5X GC buffer in the reaction
Satisfactory amplification of the products from the PCR which were of good quality as seen with Agarose gel and Sanger sequecning.
None
Easy-to-use; good for amplification of high GC content templates
Determine the annealing temperature for your product and check if you gene/target template needs High GC buffer if it has high GC content.
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