Kubo Jun, Yamanouchi Keitaro, Naito Kunihiko, Tojo Hideaki
Laboratory of Applied Genetics, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan.
J Exp Zool. 2002 Dec 1;293(7):712-8. doi: 10.1002/jez.10163.
The present paper describes the expression of a target fusion gene, WAP/hGH fused to the EGFP-expressing gene in transgenic mice derived from the transfer of transgenic embryos selected because of their expression of enhanced green fluorescent protein (EGFP). The 6.7-kb fusion gene was microinjected as a single cassette gene construct into the pronuclei of mouse zygotes. The surviving embryos were cultured and were classified according to the EGFP expression patterns at the morula or blastocyst stage. After the transfer of embryos with uniform-expression or mosaic-expression of EGFP, transgenesis occurred in 85.7% to 86% or 44.1% to 44% of the pups, respectively. No transgenic pups were derived from EGFP negative embryos. In the transgenic females, EGFP was ubiquitously expressed under the control of the CAG promoter, and hGH was expressed under the control of the WAP promoter in an appropriate fashion: hGH was secreted into the milk of lactating transgenic females. The presence or absence of the expression of EGFP coincided with that of the hGH gene in the transgenic mice. The present cassette gene construct is a useful example for circumventing the routine analyses of DNA and RNA required for the generation and maintenance of transgenic lines.
本文描述了一种目标融合基因WAP/hGH的表达情况，该融合基因与表达增强型绿色荧光蛋白（EGFP）的基因相连，存在于通过转移因表达增强型绿色荧光蛋白（EGFP）而被挑选出的转基因胚胎所获得的转基因小鼠中。将6.7kb的融合基因作为单个盒式基因构建体显微注射到小鼠受精卵的原核中。存活的胚胎进行培养，并根据桑椹胚或囊胚阶段的EGFP表达模式进行分类。在转移具有EGFP均匀表达或嵌合表达的胚胎后，转基因分别在85.7%至86%或44.1%至44%的幼崽中发生。没有从EGFP阴性胚胎获得转基因幼崽。在转基因雌性小鼠中，EGFP在CAG启动子的控制下普遍表达，而hGH在WAP启动子的控制下以适当的方式表达：hGH分泌到哺乳期转基因雌性小鼠的乳汁中。在转基因小鼠中，EGFP表达的有无与hGH基因的表达情况一致。本盒式基因构建体是一个有用的例子，可避免对转基因品系的产生和维持所需的DNA和RNA进行常规分析。
