How can the recombinant plasmid (pTYB, pTXB or pCYB vector with an insert) be analyzed by restriction digestion when the SapI or SmaI site is used for cloning?
If SapI or SmaI sites are used for cloning, the recognition sequence may not be regenerated. The KpnI site, which is located 12 nucleotides downstream of the N-terminus of the SceVMA1 intein in pTYB1-4 (or pCYB1-4 vectors), can be used for restriction analysis. The SpeI site present 17 nucleotides downstream of the N-terminus in the Mxe GyrA intein present in the pTXB vectors, can be used for restriction analysis. The NdeI (or NcoI) site may be used to cleave the 5ยด end of the insert. Other combinations of double digestions (e.g. NdeI and PstI, NcoI and PstI, XbaI and KpnI) may also be used to determine whether the clones contain the desired insert. Analysis of an insert in pTYB21, pTYB22, pTYB11 and pTYB12 can be achieved using the SacII (unique in the intein) and PstI sites..
Related Questions
- How can the recombinant plasmid (pTYB, pTXB or pCYB vector with an insert) be analyzed by restriction digestion when the SapI or SmaI site is used for cloning?
- Do the pTYB, pTXB, pTWIN or pCYB vectors carry any marker for the selection of the recombinant plasmids?
- Should a pTYB vector (or a pCYB vector) be used as a control for expression?
