SEQUENCE ANALYSIS OF TK AND HPRT GENE MUTATIONS

TK 和 HPRT 基因突变的序列分析

• 批准号: 2308734
• 负责人: MELISSA C LIECHTY
• 金额: $ 4.99万
• 依托单位: APPLIED GENETICS LABORATORIES, INC.
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-06-30 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/2308734
• 关键词: SEQUENCE; ANALYSIS; TK; HPRT; GENE

Recently developed molecular genetic techniques will be used to analyze the nature of mutagenic events occurring at the tk and hprt loci in mouse L5178Y cells exposed to chemical mutagens. Because mutation assays measure phenotype and not genotype, the possibility exists that scored mutations could result from other mechanisms. Mutant sequence analysis will determine whether the mutation assays provide true measures of mutagenic events by correlating sequence changes with mutant phenotype. Mutant molecular analyses will also allow determination of the kinds of sequence changes associated with exposure to specific chemicals and whether such changes are similar to changes known to be important in tumorigenesis. Mutants will be analyzed by Southern blotting for the presence of major deletions and/or rearrangements. Polymerase chain reaction (PCR) amplification of cDNA sequences from mutants will allow preparation of specific sequencing template without the complication of screening clone banks or preparing single-stranded sequencing template. Chemical mismatch cleavage of PCR products will be tested to further speed sequence change analysis. These streamlined techniques will allow analysis of sufficient mutants to identify trends in mutation type associated with particular chemicals. Detection of such trends will influence the specific methods used to implement Phase II, which will generally involve developing methods to further streamline mutant analysis, and to apply these methods to a test panel of mutagens. As techniques and protocols become sufficiently refined, they will be made available to other investigators as a commercial service and/or as a reagent kit.

最近开发的分子遗传技术将用于分析 在小鼠中发生在TK和HPRT基因座的诱变事件的性质 L5178Y细胞暴露于化学诱变剂。 因为突变测定 测量表型而非基因型，得分的可能性 突变可能是由其他机制引起的。 突变序列分析 将确定突变测定是否提供了真实措施 诱变事件通过将序列与突变表型的变化相关联。 突变分子分析也将允许确定类型 序列变化与暴露于特定化学物质和 这种变化是否类似于已知的变化很重要 肿瘤发生。 突变体将通过Southern印迹分析 存在重大缺失和/或重排。 聚合酶链 突变体的cDNA序列的反应（PCR）将允许 制备特定的测序模板，而没有并发症的并发症 筛选克隆库或准备单链测序模板。 PCR产品的化学不匹配裂解将进行进一步测试 速度序列变化分析。 这些简化的技术将允许 分析足够的突变体以鉴定突变类型的趋势 与特定化学物质有关。 这种趋势的检测将 影响用于实施第二阶段的特定方法，这将 通常涉及开发进一步简化突变体的方法 分析，并将这些方法应用于诱变剂的测试面板。 作为 技术和协议变得足够完善，将被制成 其他调查人员可作为商业服务和/或 试剂套件。

项目成果

Mouse chromosome-specific painting probes generated from microdissected chromosomes.

由显微解剖染色体产生的小鼠染色体特异性绘画探针。

• DOI: 10.1007/bf00352363
• 发表时间: 1995
• 期刊: Mammalian genome : official journal of the International Mammalian Genome Society
• 作者: Liechty,MC;Hall,BK;Scalzi,JM;Davis,LM;Caspary,WJ;Hozier,JC
• 通讯作者: Hozier,JC