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利用来自秀丽隐杆线虫的Tc1转座DNA元件通过P因子介导黑腹果蝇的种系转化。

P element mediated germ line transformation of Drosophila melanogaster with the Tc1 transposable DNA element from Caenorhabditis elegans.

作者信息

Szekely A A, Woodruff R C, Mahendran R

机构信息

Department of Biological Sciences, Bowling Green State University, OH 43403.

出版信息

Genome. 1994 Jun;37(3):356-66. doi: 10.1139/g94-051.

Abstract

Questions relating to the origin and regulation of mobile genetic elements are currently of considerable interest. Since it is now possible to address more precisely issues concerning the entry, dispersion, and regulation of elements within a virgin genome, one approach that may afford a better understanding of transposable elements in general could be provided by interspecific DNA transformation. Therefore, the Tc1 transposable DNA element from Caenorhabditis elegans was chosen as a proposed invading element of the Drosophila melanogaster genome. The basis for this selection resided in the inherent structural and functional similarities, as well as sequence identities, between the Caenorhabditis element and elements innate to Drosophila (e.g., P, HB1, and Uhu). Initial investigations were carried out to define a clone carrying an intact Tc1 element. This Tc1 element was inserted into a P transposon vector and two P-Tc1-ry+ constructs, differing only in insert orientation, were identified. P element mediated germ line transfer was then used to generate a transformant that was genetically and molecularly identified as containing a single, structurally intact Tc1 element at cytological location 64C4-5 on the third chromosome. The single P[(Tc1,ry+)]SAS-B insertion was thereafter mobilized by using a P[ry+ delta 2-3] element as a transposase source, and the genetic and molecular data suggested that the insertion had been successfully reintegrated to a variety of genomic locations. On the basis of genetic and molecular analyses, the Tc1 element in the P[Tc1,ry+)] transformed stock is not highly unstable in germ line and somatic tissues.

摘要

目前,与移动遗传元件的起源和调控相关的问题备受关注。由于现在能够更精确地解决有关元件在原始基因组中的进入、扩散和调控问题,种间DNA转化可能为更全面地理解转座元件提供一种方法。因此,选择了秀丽隐杆线虫的Tc1转座DNA元件作为拟侵入黑腹果蝇基因组的元件。做出这一选择的依据是秀丽隐杆线虫元件与果蝇固有元件(如P、HB1和Uhu)之间在结构和功能上的内在相似性以及序列同源性。最初进行了研究以确定携带完整Tc1元件的克隆。将该Tc1元件插入P转座子载体中,并鉴定出两个仅在插入方向上不同的P-Tc1-ry+构建体。然后利用P元件介导的种系转移产生一个转化体,通过遗传和分子鉴定,该转化体在第三条染色体的细胞学位置64C4-5处含有单个结构完整的Tc1元件。此后,利用P[ry+ delta 2-3]元件作为转座酶来源,使单个P[(Tc1,ry+)]SAS-B插入发生移动,遗传和分子数据表明该插入已成功重新整合到各种基因组位置。基于遗传和分子分析,P[Tc1,ry+)]转化品系中的Tc1元件在种系和体细胞组织中并非高度不稳定。

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