来自流式分选染色体的DNA序列文库的构建与分析：实践与理论考量
Construction and analysis of DNA sequence libraries from flow-sorted chromosomes: practical and theoretical considerations.
作者信息
Griffith J K, Cram L S, Crawford B D, Jackson P J, Schilling J, Schimke R T, Walters R A, Wilder M E, Jett J H
出版信息
Nucleic Acids Res. 1984 May 11;12(9):4019-34. doi: 10.1093/nar/12.9.4019.
We describe the construction and analysis of recombinant DNA libraries representative of chromosomes 1 and 2 of Chinese hamster (Cricetulus griseus). Propidium-iodide stained chromosomes were purified by flow cytometric analysis and sorting, and EcoRI digests of purified DNA were cloned into the bacteriophage vector Charon 4A. These libraries contain DNA complementary to 63% and 69% of nick-translated DNA derived from flow-purified chromosomes 1 and 2, respectively. However, sequences complementary to only 24% and 35% of a total Chinese hamster genomic DNA tracer were hybridized in parallel renaturation experiments. The chromosome 2 library contained DNA sequences encoding dihydrofolate reductase (dhfr), a gene previously mapped to Chinese hamster chromosome 2. No sequences complementary to dhfr were found in the library constructed from chromosome 1 DNA. These analyses are discussed with regard to the current limitations and future strategies for the construction of chromosome-specific DNA sequence libraries of high purity and completeness.
我们描述了代表中国仓鼠（黑线仓鼠）1号和2号染色体的重组DNA文库的构建及分析。用碘化丙啶染色的染色体通过流式细胞术分析和分选进行纯化，纯化后的DNA经EcoRI酶切后克隆到噬菌体载体Charon 4A中。这些文库分别包含与来自流式纯化的1号和2号染色体的63%和69%的缺口平移DNA互补的DNA。然而，在平行复性实验中，仅与中国仓鼠基因组DNA示踪剂总量的24%和35%互补的序列发生了杂交。2号染色体文库包含编码二氢叶酸还原酶（dhfr）的DNA序列，该基因先前已定位到中国仓鼠2号染色体上。在由1号染色体DNA构建的文库中未发现与dhfr互补的序列。针对目前构建高纯度和完整性的染色体特异性DNA序列文库的局限性及未来策略，对这些分析进行了讨论。
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