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dc.contributor.authorChaudhry, M Ahmaden
dc.contributor.authorFox, Margareten
dc.date.accessioned2010-08-18T13:44:41Z
dc.date.available2010-08-18T13:44:41Z
dc.date.issued1990-09
dc.identifier.citationMethylmethane-sulphonate and X-ray-induced mutations in the Chinese hamster hprt gene: mRNA phenotyping using polymerase chain reactions. 1990, 5 (5):497-504 Mutagenesisen
dc.identifier.issn0267-8357
dc.identifier.pmid2263207
dc.identifier.doi10.1093/mutage/5.5.497
dc.identifier.urihttp://hdl.handle.net/10541/109874
dc.description.abstractAlterations in the hprt gene of Chinese hamster cells were determined in 71 spontaneous, methylmethane sulphonate (MMS)- and X-ray-induced mutants, using the Southern blot hybridization technique. Among 41 MMS-induced mutants, deletions eliminating the whole gene were observed in 17 cases (41%). Analysis of 20 X-ray-induced mutants revealed the presence of similar deletions in nine of them (45%). No evidence of deletion was found in 10 spontaneous mutants. To investigate the possibility of small deletions, 18 MMS-induced mutants were studied with probes derived from exons 3 and 9 but no evidence of specific deletion of these two exons was found. The polymerase chain reaction (PCR) was used to phenotype hprt transcripts in 48 MMS, X-ray and spontaneous Chinese hamster mutants by amplifying the coding region of their cDNA. Among 22 MMS-induced mutants the message was present in 16 instances; 11 had normal levels while three had much lower levels of transcription. A further two mutants had mRNA of reduced size as revealed by the use of primers for PCR 3' to those routinely used. An analysis of 20 X-ray-induced mutants showed the presence of hprt mRNA in 11 of them with five having low levels of transcription. Among six spontaneous mutants, four were negative for mRNA on standard Northern blots and in one the message was only detected after PCR amplification. Direct DNA sequencing of 10 mutants revealed the presence of base substitutions in five of them while a 7 bp deletion was found in another. No mutations were found in another four mutants, suggesting the presence of mutation outside the coding region.
dc.language.isoenen
dc.subject.meshAnimals
dc.subject.meshBase Sequence
dc.subject.meshBlotting, Northern
dc.subject.meshBlotting, Southern
dc.subject.meshCell Line
dc.subject.meshExons
dc.subject.meshGenes
dc.subject.meshHypoxanthine Phosphoribosyltransferase
dc.subject.meshMethyl Methanesulfonate
dc.subject.meshMolecular Sequence Data
dc.subject.meshMutation
dc.subject.meshPhenotype
dc.subject.meshPolymerase Chain Reaction
dc.subject.meshRNA, Messenger
dc.titleMethylmethane-sulphonate and X-ray-induced mutations in the Chinese hamster hprt gene: mRNA phenotyping using polymerase chain reactions.en
dc.typeArticleen
dc.contributor.departmentCRC Department of Biochemical Genetics, Paterson Institute for Cancer Research, Christie Hospital and Holt Radium Institute, Manchester, UK.en
dc.identifier.journalMutagenesisen
html.description.abstractAlterations in the hprt gene of Chinese hamster cells were determined in 71 spontaneous, methylmethane sulphonate (MMS)- and X-ray-induced mutants, using the Southern blot hybridization technique. Among 41 MMS-induced mutants, deletions eliminating the whole gene were observed in 17 cases (41%). Analysis of 20 X-ray-induced mutants revealed the presence of similar deletions in nine of them (45%). No evidence of deletion was found in 10 spontaneous mutants. To investigate the possibility of small deletions, 18 MMS-induced mutants were studied with probes derived from exons 3 and 9 but no evidence of specific deletion of these two exons was found. The polymerase chain reaction (PCR) was used to phenotype hprt transcripts in 48 MMS, X-ray and spontaneous Chinese hamster mutants by amplifying the coding region of their cDNA. Among 22 MMS-induced mutants the message was present in 16 instances; 11 had normal levels while three had much lower levels of transcription. A further two mutants had mRNA of reduced size as revealed by the use of primers for PCR 3' to those routinely used. An analysis of 20 X-ray-induced mutants showed the presence of hprt mRNA in 11 of them with five having low levels of transcription. Among six spontaneous mutants, four were negative for mRNA on standard Northern blots and in one the message was only detected after PCR amplification. Direct DNA sequencing of 10 mutants revealed the presence of base substitutions in five of them while a 7 bp deletion was found in another. No mutations were found in another four mutants, suggesting the presence of mutation outside the coding region.


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