Molecular analysis of complex human cell populations: mutational spectra of MNNG and ICR-191.
We describe a method to identify and enumerate mutants at the nucleotide level in complex cell populations. Several thousand different mutants were induced at the HPRT locus in human lymphoblastoid cultures by either MNNG, an alkylating agent, or by ICR-191, a substituted acridine. HPRT mutants were selected en masse by resistance to 6-thioguanine. The most frequent mutations (hotspots) in HPRT exon 3 were determined by a combination of denaturing gradient gel electrophoresis and polymerase chain reaction. MNNG predominantly produced GC----AT transitions at nucleotides in a GGGGGG sequence, while ICR-191 produced both +1 frameshifts in the same GGGGGG sequence and +1 frameshifts in a CCC sequence.
Cariello
N F
NF
Massachusetts Institute of Technology, Center for Environmental Health Sciences, Cambridge 02139.
Keohavong
P
P
Kat
A G
AG
Thilly
W G
WG
D000585:Aminacrine / Q000031:analogs & derivatives / Q000633:toxicity*; D000609:Aminoacridines / Q000633:toxicity*; D001402:B-Lymphocytes; D001483:Base Sequence; D002460:Cell Line; D003001:Cloning, Molecular; D004586:Electrophoresis; D005091:Exons; D006801:Humans; D007041:Hypoxanthine Phosphoribosyltransferase / Q000235:genetics*; D008769:Methylnitronitrosoguanidine / Q000633:toxicity*; D008969:Molecular Sequence Data; D009154:Mutation; D009588:Nitrogen Mustard Compounds / Q000633:toxicity*; D016133:Polymerase Chain Reaction
D000609:Aminoacridines; D009588:Nitrogen Mustard Compounds; D008769:Methylnitronitrosoguanidine; D000585:Aminacrine; C004294:acridine half-mustard; D007041:Hypoxanthine Phosphoribosyltransferase