Generation of transgenic rats using YAC and BAC DNA constructs.
Transgenic rats with a simple plasmid vector smaller than 20 Kb show insufficient expression and tissue specificity of the introduced transgene. Vectors derived from yeast artificial chromosome (YAC) and bacterial artificial chromosome (BAC), consisting of DNA fragments up to approximately 1 Mb (YAC) and approximately 200 Kb (BAC), respectively, and containing various endogenous regulatory sequences, were expected to work well and showed expression profiles comparable to their endogenous counterparts in transgenic animals. While attempting to make transgenic rats using YAC and BAC vectors, we faced two problems: how to prepare sufficiently concentrated intact DNA and how to reliably microinject a large DNA fragment into the fragile pronuclear ova of the rat. After solving these problems, we were able to make transgenic rats by introducing YAC/BAC gene constructs (YACs/BACs) into the pronuclear ova. And then we examined the relative transcription rates of these genes in the transgenic rats. In this chapter, we focus on this injection process.
Takahashi
Ri-Ichi
R
Phoenixbio Co. Ltd., Tochigi, Japan.
Ueda
Masatsugu
M
D000818:Animals; D022202:Chromosomes, Artificial, Bacterial / Q000235:genetics*; D018244:Chromosomes, Artificial, Yeast / Q000235:genetics*; D004247:DNA / Q000008:administration & dosage* / Q000235:genetics; D004867:Equipment Design; D005822:Genetic Vectors; D008845:Microinjections / Q000295:instrumentation / Q000379:methods*; D010063:Ovum / Q000166:cytology; D051381:Rats; D055647:Rats, Transgenic / Q000235:genetics*; D019076:Transgenes
D004247:DNA