Positioning isolation and biochemical analysis of single DNA molecules based on nanomanipulation and single-molecule PCR.
Recently, the isolation and biochemical analysis of DNA at the single-molecule level has been recognized as very important for genetic research and clinical analysis. A unique technique for the positioning, dissection, and isolation of single DNA molecules using atomic force microscopy (AFM) has been demonstrated. Full-length genome DNA molecules were first deposited and stretched by a modified "molecular combing" technique onto a 3-aminopropyl triethoxysilane-coated mica substrate. A single DNA fragment was dissected from one of those genome DNA strands with the AFM tip at the desired position, and then isolated (or picked up) after a special operation called "kneading". All the operations including imaging, dissection, and isolation could be carried out with one tip. The isolated DNA fragment on the AFM tip could be successfully amplified by single-molecule PCR.
Lü
Jun-Hong
JH
Shanghai Institute of Applied Physics, Chinese Academy of Sciences, Shanghai 201800, China, and Bio-X Life Science Research Center, Shanghai JiaoTong University, Shanghai 200030, China.
Li
Hai-Kuo
HK
An
Hong-Jie
HJ
Wang
Guo-Hua
GH
Wang
Ying
Y
Li
Min-Qian
MQ
Zhang
Yi
Y
Hu
Jun
J
D004247:DNA / Q000032:analysis* / Q000302:isolation & purification*; D018625:Microscopy, Atomic Force; D036103:Nanotechnology / Q000379:methods*; D010957:Plasmids / Q000032:analysis / Q000302:isolation & purification; D016133:Polymerase Chain Reaction / Q000379:methods*
D004247:DNA