[Use PCR synthesis large fragment DNA].
Synthesis Large Fragment DNA using PCR (SLFD PCR) is a useful method to synthesize large fragment DNA. Use a known 500 approximately 600 basepair DNA fragment as PCR template, a series of 5' terminal primers are designed, and these primers are overlap one by one from 5' terminal to 3' terminal, the net DNA is just what you want to synthesize. The last 3' terminal primer of these primers has a BamH I site, and behind the BamH I site there are 15 bp overlap the 5' terminal of the template. Another downstream primer complement the 3' terminal of the template, and has a BamH I site too. PCR begin using the innerest 5' terminal primer and the downstream primer. After 10 cycles of PCR, use the product of the PCR as the template of next round PCR, but this time the upstream primer change to the 5' terminal outer primer . So do the PCRs, till all the 5' terminal primers take part in the PCR. Clone the final PCR product and BamH I cut the original DNA template, The DNA synthesis complete. It's a useful method to synthesize 100 approximately 200 bp DNA fragment, even more long DNA fragment.
Li
Wei-Dong
WD
Wuhan Institute of Virology, Chinese Academy of Science, Wuhan 430071, China. wd_bio@sohu.com
Liang
Bu-Feng
BF
Qi
Zi-Bai
ZB
D023181:Antimicrobial Cationic Peptides / Q000235:genetics*; D001483:Base Sequence; D004247:DNA / Q000096:biosynthesis*; D053938:DNA Fragmentation; D017931:DNA Primers; D050939:Gene Fusion; D015696:Gene Products, tat / Q000235:genetics* / Q000302:isolation & purification; D006678:HIV / Q000737:chemistry; D008555:Melitten / Q000235:genetics; D010957:Plasmids; D016133:Polymerase Chain Reaction / Q000379:methods*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics; D054322:tat Gene Products, Human Immunodeficiency Virus
D023181:Antimicrobial Cationic Peptides; D017931:DNA Primers; D015696:Gene Products, tat; D011993:Recombinant Fusion Proteins; D054322:tat Gene Products, Human Immunodeficiency Virus; D008555:Melitten; C031161:cecropin A; D004247:DNA