Identification of expressed genes by laser-mediated manipulation of single cells.
We describe a rapid noncontact method for the capture of single cells or small tissue areas of any size or shape directly within the cap of a common microfuge tube. Prior to the laser-mediated transfer, the specimen is isolated by laser microbeam microdissection, forming a clear-cut gap around the selected area. Laser treatment does not impair subsequent RNA analysis. We have used this method to isolate a single cell from archival colon adenocarcinoma, and were able to detect point mutations within codon 12 of c-Ki-ras2 mRNA after nested RT-PCR analysis.
Laser Laboratory and Molecular Biology, First Medical Department, Academic Hospital München-Harlaching, Germany.
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Schütze
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Lahr
D000230:Adenocarcinoma / Q000235:genetics / Q000473:pathology*; D002469:Cell Separation / Q000379:methods*; D003062:Codon / Q000235:genetics; D003110:Colonic Neoplasms / Q000235:genetics / Q000473:pathology*; D019003:Deoxyribonuclease HpaII; D015252:Deoxyribonucleases, Type II Site-Specific; D011905:Genes, ras; D006801:Humans; D007834:Lasers; D008867:Microtomy / Q000379:methods; D017354:Point Mutation; D016133:Polymerase Chain Reaction / Q000379:methods; D012333:RNA, Messenger / Q000235:genetics
D003062:Codon; D012333:RNA, Messenger; D019003:Deoxyribonuclease HpaII; D015252:Deoxyribonucleases, Type II Site-Specific; C027046:TCGA-specific type II deoxyribonucleases