Tilghman S M, Tiemeier D C, Polsky F, Edgell M H, Seidman J G, Leder A, Enquist L W, Norman B, Leder P
Proc Natl Acad Sci U S A. 1977 Oct;74(10):4406-10. doi: 10.1073/pnas.74.10.4406.
We have developed a general approach to the cloning of specific segments of the mammalian genome that involves a two-step purification of EcoRI fragments of mammalian DNA and their in vitro insertion into a suitably constructed EK2 derivative of bacteriophage lambda. The combination of fragment purification, exclusion of parental-type recombinants, and simple phage screening techniques permits the isolation of virtually any gene segment for which there is an identifying hybridization probe. We illustrate the approach by describing the cloning of an approximately 7000-base-long segment of mouse DNA containing globin and surrounding gene sequences.
我们已经开发出一种克隆哺乳动物基因组特定片段的通用方法,该方法包括对哺乳动物DNA的EcoRI片段进行两步纯化,并将其体外插入到经过适当构建的噬菌体λ的EK2衍生物中。片段纯化、排除亲本型重组体以及简单的噬菌体筛选技术相结合,使得几乎任何有识别性杂交探针的基因片段都能被分离出来。我们通过描述克隆一段约7000个碱基长的小鼠DNA片段(包含珠蛋白及周围基因序列)来说明该方法。