ALU POLYMERASE CHAIN-REACTION - A METHOD FOR RAPID ISOLATION OF HUMAN-SPECIFIC SEQUENCES FROM COMPLEX DNA SOURCES
ALU POLYMERASE CHAIN-REACTION - A METHOD FOR RAPID ISOLATION OF HUMAN-SPECIFIC SEQUENCES FROM COMPLEX DNA SOURCES
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DOI：
10.1073/pnas.86.17.6686
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发表时间：
1989-09-01
影响因子：
11.1
通讯作者：
CASKEY, CT
中科院分区：
文献类型：
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作者：
NELSON, DL;LEDBETTER, SA;CASKEY, CT
目前绘制人类基因组图谱的努力集中在单个染色体或更小的区域，并且经常依赖于体细胞杂交的使用。我们报道了应用聚合酶链式反应直接从啮齿动物细胞背景中含有人类基因组区域的杂交细胞中扩增人类DNA，方法是使用针对人类Alu重复元件的引物。我们从杂合细胞和克隆的DNA中进行了人HPRT基因片段的Alu定向扩增，并通过序列分析鉴定了参与这种扩增的Alu重复序列。我们还展示了该技术在确定酵母人工染色体中克隆的人类X染色体大片段的染色体位置的应用，以及该方法在从克隆的人类序列中制备DNA探针方面的普遍适用性。该技术可以快速绘制基因图谱，并为分离和分析特定的染色体区域提供了一种简单的方法。
Current efforts to map the human genome are focused on individual chromosomes or smaller regions and frequently rely on the use of somatic cell hybrids. We report the application of the polymerase chain reaction to direct amplification of human DNA from hybrid cells containing regions of the human genome in rodent cell backgrounds using primers directed to the human Alu repeat element . We demonstrate Alu-directed amplification of a fragment of the human HPRT gene from both hybrid cell and cloned DNA and identify through sequence analysis the Alu repeats involved in this amplification. We also demonstrate the application of this technique to identify the chromosomal locations of large fragments of the human X chromosome cloned in a yeast artificial chromosome and the general applicability of the method to the preparation of DNA probes from cloned human sequences. The technique allows rapid gene mapping and provides a simple method for the isolation and analysis of specific chromosomal regions.