Synthesis of nucleic acid probes on membrane supports: a procedure for the removal of unincorporated precursors.
We have used DNA bound to small pieces of nylon membrane for the synthesis of radioactive probes. The DNA to be used for generating the probe(s) is first bound to nylon membranes and then introduced into the reaction mix. The labeling reaction takes place on the membrane and therefore allows easy removal of unincorporated precursors by simple washing for 1-2 min. The clean labeled probe is eluted from the membrane in formamide or in water and is ready for use. This DNA-membrane can be stored for reuse. Synthesis of probes on a solid support such as nylon membrane thus circumvents problems associated with chromatographic manipulations needed for the separation of labeled DNA from unicorporated precursors. Probes synthesized in this manner are as efficient in detecting nucleic acid sequences as those synthesized in solution.
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Jules Stein Eye Institute, University of California School of Medicine, Los Angeles 90024-1771.
D015152:Blotting, Northern; D015139:Blotting, Southern; D015342:DNA Probes / Q000138:chemical synthesis*; D003845:Deoxycytosine Nucleotides / Q000378:metabolism; D008567:Membranes, Artificial; D008722:Methods; D009693:Nucleic Acid Hybridization; D009757:Nylons; D010761:Phosphorus Radioisotopes
D015342:DNA Probes; D003845:Deoxycytosine Nucleotides; D008567:Membranes, Artificial; D009757:Nylons; D010761:Phosphorus Radioisotopes; C024107:2'-deoxycytidine 5'-triphosphate