An improved nuclear extract preparation method.
A rapid, efficient, and highly reproducible procedure for nuclear extract preparation is described. The method uses lysolecithin (lysophosphatidylcholine) to disrupt plasma membranes and requires no detergents or douncing. Soluble extracts prepared by this method are comparable to conventional nuclear extracts in all assays tested. Lysolecithin nuclear extracts are competent for RNA polymerase II and III transcription, DNA replication, pre-mRNA splicing, and sequence specific DNA-protein binding. Nuclear extracts can be prepared on a small scale (10(7) cells) as well as for preparative purposes by this method.
Zerivitz
K
K
Department of Microbial Genetics, Karolinska Institute, Stockholm, Sweden.
Akusjärvi
G
G
D000256:Adenoviridae / Q000235:genetics; D002467:Cell Nucleus / Q000032:analysis* / Q000502:physiology; D004261:DNA Replication; D004279:DNA, Viral / Q000096:biosynthesis; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism; D012321:DNA-Directed RNA Polymerases / Q000235:genetics; D006367:HeLa Cells; D006801:Humans; D008244:Lysophosphatidylcholines; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000096:biosynthesis / Q000502:physiology; D013539:Simian virus 40 / Q000235:genetics; D013347:Subcellular Fractions; D014158:Transcription, Genetic
D004279:DNA, Viral; D004268:DNA-Binding Proteins; D008244:Lysophosphatidylcholines; D012333:RNA, Messenger; D012321:DNA-Directed RNA Polymerases