hi,
i am designing a synthetic gene and i need to have Sal1and EcoRV restriction site at the 5' and 3' terminals.can i just insert the restriciton site of the enzyme on both ends and then design the whole gene..or do i need any additional base pairs in addition to these restriction sites on 5' and 3' ends....
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2 replies to this topic
#2
phage434
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Posted 10 April 2009 - 03:57 AM
You need additional bases. I would add at least 6, preferably 8 bases 5' of your cut site. I would suggest switching to a different enzyme than SalI, which has a very poor reputation for cutting PCR fragments.
#3
perneseblue
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Unlimited ligation works!
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Posted 10 April 2009 - 10:05 PM
phage434, on Apr 10 2009, 03:57 AM, said:
I would suggest switching to a different enzyme than SalI, which has a very poor reputation for cutting PCR fragments.
XhoI and SalI form compatible overhangs
May your PCR products be long, your protocols short and your boss on holiday
