Ion pair-reversed phase HPLC approach facilitates subcloning of PCR products and screening of recombinant colonies.
The isolation of a single DNA molecule for cloning is technically difficult, and the subsequent screening of colonies for recombinant DNA clones is time consuming. Ion pair-reversed phase HPLC (IP-RP-HPLC) analysis of nucleic acids improves the resolution and isolation of PCR products to be cloned. We demonstrate that PCR products analyzed and collected using the IP-RP-HPLC approach (WAVE DNA Fragment Analysis System) can be cloned directly into a plasmid vector. In addition, we demonstrate that when IP-RP-HPLC analysis is extended to the colony screening process, the time required for these procedures is reduced.
Shaw-Bruha
C M
CM
Transgenomic Inc., Omaha, NE, USA. cshaw@transgenomic.com
Lamb
K A
KA
D002851:Chromatography, High Pressure Liquid / Q000191:economics / Q000379:methods*; D003001:Cloning, Molecular / Q000379:methods*; D004247:DNA / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification*; D010957:Plasmids / Q000737:chemistry / Q000235:genetics; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000032:analysis; D013997:Time Factors
D011994:Recombinant Proteins; D004247:DNA