An efficient and site-specific gene trimming method.
Trimming a DNA strand into a precisely determined fragment can be carried out efficiently by an improved method involving a site-specific trim-primer and a single-stranded DNA template which is generated from a multifunctional vector, pTZ18R, and linearized by using an Eco RI-pTZ18R splinter. A complementary DNA strand is synthesized by DNA polymerase I (Klenow), and the 3'-end of the template upstream from the annealed primer is trimmed by subsequent T4 DNA polymerase reaction. An ATG translation initiator codon or a termination codon can be incorporated into the trim-primer, providing versatility to this single-stranded DNA-initiated gene trimming method that can be applied to subcloning and expression of any DNA fragment with known terminal sequences.
Eun
H M
HM
University of Calgary.
Kang
Y
Y
Kang
S M
SM
Bae
Y S
YS
Yoon
J W
JW
D001709:Biotechnology; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics* / Q000302:isolation & purification; D004256:DNA Polymerase I; D004260:DNA Repair; D004277:DNA, Single-Stranded / Q000235:genetics; D005821:Genetic Techniques; D006801:Humans; D007328:Insulin; D011384:Proinsulin / Q000235:genetics; D011498:Protein Precursors / Q000235:genetics
D004277:DNA, Single-Stranded; D007328:Insulin; D011498:Protein Precursors; C013737:preproinsulin; D004247:DNA; D011384:Proinsulin; D004256:DNA Polymerase I