New answers tagged cloning
The error rate of PCR is still very high in comparison to natural bacteria DNA replication of its plasmids. Natural bacteria DNA replication has an error rate of approximately 1 in 10 billion, and the best PCR polymerase commercially available (Q5 from NEB) has an error rate of approximately 1 in 1 million. Therefore, cloning fragments into bacteria and ...
You can use PCR products in Sanger sequencing; it is very common. Using PCR products instead of cloned genes does raise a set of problems that are less of a concern than with cloned sequences, such as the presence of incomplete or incorrect PCR products, but there are standard and simple solutions for most of these concerns.
From my experience I would say that it is no problem to freeze the digested DNA, when the enzyme is inactivated (even when not, for this time period I wouldn't expect much damage). However, if possible, I would rather opt to freeze the PCR product and make the digest fresh, since the overhanging ends are kind of sensitive to hydrolytic degradation. I have ...
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