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The mutagenic specificity of ultravi...

Drobetsky, Elliot.

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The mutagenic specificity of ultraviolet light at the aprt locus in Chinese hamster ovary cells.

Record Type:	Electronic resources : Monograph/item
Title/Author:	The mutagenic specificity of ultraviolet light at the aprt locus in Chinese hamster ovary cells./
Author:	Drobetsky, Elliot.
Published:	Ann Arbor : ProQuest Dissertations & Theses, : 1990,
Description:	166 p.
Notes:	Source: Dissertation Abstracts International, Volume: 52-11, Section: B, page: 5665.
Contained By:	Dissertation Abstracts International52-11B.
Subject:	Molecular biology. -
Online resource:	http://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=NN60963
ISBN:	9780315609631

The mutagenic specificity of ultraviolet light at the aprt locus in Chinese hamster ovary cells.
Drobetsky, Elliot.

The mutagenic specificity of ultraviolet light at the aprt locus in Chinese hamster ovary cells. - Ann Arbor : ProQuest Dissertations & Theses, 1990 - 166 p.

Source: Dissertation Abstracts International, Volume: 52-11, Section: B, page: 5665.

Thesis (Ph.D.)--York University (Canada), 1990.

In order to increase our understanding of fundamental mechanisms of mutagenesis in mammalian cells, we have developed a method for studying mutational specificity at the endogenous aprt locus in CHO cells. This approach, based on the in vivo recombinational rescue of mutant alleles, has been used to clone and sequence the coding region of 34 UV-induced aprt mutants. Among the mutants recovered, 26 were single base substitutions including 17 G:C$\to$A:T transitions and a single A:T$\to$G:C transition. Three of the four possible transversions accounted for the remaining eight mutations. The G:C$\to$T:A transversion was not recovered. Six tandem double or closely neighbouring (non-tandem) double base substitutions, one double mutation consisting of a G:C$\to$T:A transversion and an adjacent frameshift, as well as one single frameshift mutation were also recovered. UV-induced mutation appears to be targeted to dipyrimidine sites with only two exceptions. These include two double mutations where only one of the base substitutions occurred at a dipyrimidine site. The observed specificity of UV-light induced mutations at the aprt locus is consistent with the notion that G:C$\to$A:T transitions result primarily from the (6-4) pyrimidine-pyrimidone lesion.

ISBN: 9780315609631Subjects--Topical Terms:

517296
Molecular biology.

The mutagenic specificity of ultraviolet light at the aprt locus in Chinese hamster ovary cells.
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100 1 $a Drobetsky, Elliot. $3 3427650
245 1 4 $a The mutagenic specificity of ultraviolet light at the aprt locus in Chinese hamster ovary cells.
260 1 $a Ann Arbor : $b ProQuest Dissertations & Theses, $c 1990
300 $a 166 p.
500 $a Source: Dissertation Abstracts International, Volume: 52-11, Section: B, page: 5665.
500 $a Supervisor: G. W. Glickman.
502 $a Thesis (Ph.D.)--York University (Canada), 1990.
520 $a In order to increase our understanding of fundamental mechanisms of mutagenesis in mammalian cells, we have developed a method for studying mutational specificity at the endogenous aprt locus in CHO cells. This approach, based on the in vivo recombinational rescue of mutant alleles, has been used to clone and sequence the coding region of 34 UV-induced aprt mutants. Among the mutants recovered, 26 were single base substitutions including 17 G:C$\to$A:T transitions and a single A:T$\to$G:C transition. Three of the four possible transversions accounted for the remaining eight mutations. The G:C$\to$T:A transversion was not recovered. Six tandem double or closely neighbouring (non-tandem) double base substitutions, one double mutation consisting of a G:C$\to$T:A transversion and an adjacent frameshift, as well as one single frameshift mutation were also recovered. UV-induced mutation appears to be targeted to dipyrimidine sites with only two exceptions. These include two double mutations where only one of the base substitutions occurred at a dipyrimidine site. The observed specificity of UV-light induced mutations at the aprt locus is consistent with the notion that G:C$\to$A:T transitions result primarily from the (6-4) pyrimidine-pyrimidone lesion.
520 $a The extent to which the cellular processing of shuttle vector-carried genes reflects that of endogenous chromosomal loci has been a subject of considerable controversy. In order to address this issue, we have developed a retroviral-based shuttle vector carrying the CHO aprt gene stably integrated into the genome, for studying mutational specificity in mammalian cells. Initially, we have characterized a collection of UV-induced mutants in a CHO cell background. We have therefore been able to directly compare this shuttle vector data to that previously obtained for UV-induced mutation at the endogenous CHO aprt locus. Although some potential differences between the two spectra have been noted, there appears to be a remarkable similarity in the distribution and site specificity of UV-induced mutations. These similarities extend to extrachromosomal shuttle vectors as well, and consolidate the role of shuttle vectors as powerful analytical tools for studying mechanisms of point mutagenesis in mammalian cells.
520 $a Previous studies of mutational specificity in mammalian cells have emphasized either hemizygous endogenous genes, or genetic targets carried on shuttle vectors. However, it has been demonstrated that loss of heterozygosity plays a major role in the expression of recessive phenotypes at autosomal loci. In order to address this issue, we have developed a system for studying mutational specificity at the heterozygous aprt locus in CHO cells. The strategy employed is based on restriction fragment analysis and DNA sequencing of enzymatically amplified mutant aprt alleles. We have demonstrated the usefulness of this approach through the characterization of a collection of aprt mutants with respect to the role played by loss of heterozygosity in ultraviolet light-induced mutagenesis. It was concluded that LOH does not play an important role in the expression of the APRT phenotype in CHO cells following exposure to UV light. We have also demonstrated that restriction fragment analysis of PCR-amplified mutant aprt alleles can be used to shed light on the identity of the premutational lesion responsible for a UV-induced mutational hotspot at the aprt locus.
590 $a School code: 0267.
650 4 $a Molecular biology. $3 517296
650 4 $a Cellular biology. $3 3172791
690 $a 0307
690 $a 0379
710 2 $a York University (Canada). $3 1017889
773 0 $t Dissertation Abstracts International $g 52-11B.
790 $a 0267
791 $a Ph.D.
792 $a 1990
793 $a English
856 4 0 $u http://pqdd.sinica.edu.tw/twdaoapp/servlet/advanced?query=NN60963