gene expression

Cheng Luo cheng.luo at
Fri Oct 24 17:34:34 EST 1997

Xiao-hua Xue wrote:
> Hi,everybody,
>   I have been trying to express a gene from cDNA library using xenopus
> oocytes and it hasn't got any expression yet after I tried twice. There is
> about 200 nucleotides at 5'-terminal before the start codon. Could this
> (long non-coding sequence) be one of the reasons for no expression?
>   Any help will be really appreciated.
> Regards,
> Hua

It may affect. The best way is PCR-cloning, you design a primer around
the start codon, just add the cloning RE site in the 5'-end of the
primer, or bury the RT site in the template sequence in the same frame
order as start codon. Usually NcoI and NdeI can be just buried in the
start codon, of course you can also use other cloning enzyme if you keep
the frame.

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