Have you seen these papers: M Geiser, R Cebe, D Drewello, and R Schmitz. Integration of pcr fragments at any specific site within cloning vectors without the use of restriction enzymes and dna ligase. Biotechniques, 31(1):88–90, 2001.
W Wang and B A Malcolm. Two-stage pcr protocol allowing introduction of multiple mutations, deletions and insertions using quikchange site-directed mutagenesis. Biotechniques, 26(4):680–682, 1999. If i recall correctly, Geiser el al inserted a >1kb fragment with a modified Quickchange method. On Mon, Dec 1, 2008 at 12:54 PM, Raji Edayathumangalam <[EMAIL PROTECTED]> wrote: > Hi Folks, > > Sorry for the non-xtallo posting. > > I am curious to hear what is the longest insert anyone has cloned using a > modification of the Quikchange cloning strategy. Basically, > ligation-independent cloning by strapping on homologous regions of the > vector onto the primers which also generate the initial PCR product. I plan > to proceed with my insert which is ~ 2kb and am curious to get some feedback > if you have successfully cloned inserts > 1.5kb using the above strategy. > > Many thanks. > Raji > Michael Giffin The Scripps Research Institute Department of Molecular and Experimental Medicine 10550 North Torrey Pines Road, MEM-131 La Jolla, CA 92037 email: [EMAIL PROTECTED] lab: 858-784-7758
