Cloning big fragments

LoPony ctfrench at wwisp.com
Tue May 30 23:59:30 EST 2000


Hi Greg -

Is your plasmid dephosphorylated?  Try using more insert...


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Christopher Todd French
UAB Dept. of Comparative Medicine
1670 Univ. Blvd, VH 422
1530 3rd Ave. S
Birmingham, AL 35294-0019

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""ARNDT, Greg (JJRAU)"" <garndt at MEDAU.JNJ.COM> wrote in message
news:9F493097D6CCD211B18F0008C7FA88CEA09FC2 at JNJAUSYEXS1...
> Hello,
>
> We have had a continuous problem in cloning large PCR fragments (5-8kb)
into a smaller cloning vector (3-5kb).  We have used various restriction
enzyme site ends.  Does anyone have any helpful hints/tricks for cloning
5-8kb PCR or non-PCR DNA fragments into conventional cloning sites in 3-5kb
vectors?
>
> Regards
>
> Dr Greg Arndt
> Senior Research Scientist, Project Leader
> Johnson & Johnson Research
> Level 4, 1 Central Avenue
> Eveleigh, NSW 1430
> Sydney, Australia
> Tel: 61-2-8396-5837
> Fax: 61-2-8396-5811
> Email: garndt at medau.jnj.com
> Mailing Address:
> Locked Bag 4555
> Strawberry Hills, NSW 2012
> ---







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