Efficient and Specific Ribozyme-Mediated Reduction of Bovine α -Lactalbumin Expression in Double Transgenic Mice

• Published in: Proceedings of the National Academy of Sciences - PNAS Vol. 93; no. 13; pp. 6698 - 6703
• Main Authors: L'Huillier, Phil J., Soulier, Solange, Stinnakre, Marie-Georges, Lepourry, Laurence, Davis, Stephen R., Mercier, Jean-Claude, Vilotte, Jean-Luc
• Format: Journal Article
• Language: English
• Published: United States National Academy of Sciences of the United States of America 25.06.1996; National Acad Sciences; National Academy of Sciences
• Subjects: Animals; Base Sequence; Cattle; Cellular Biology; DNA; Gels; Genes; Lactalbumin - analysis; Lactalbumin - genetics; Life Sciences; Mammary glands; Messenger RNA; Mice; Mice, Transgenic; Milk - chemistry; Molecular Sequence Data; Molecules; Oligodeoxyribonucleotides; Ribonucleic acid; RNA; RNA, Catalytic - metabolism; RNA, Messenger - genetics; RNA, Messenger - metabolism; Rodents; Transgenes; Transgenic animals; Ungulates; Untranslated regions
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.93.13.6698

Transgenic mice carrying a bovine α -lactal-bumin (α -lac) specific ribozyme gene under the transcriptional control of the mouse mammary tumor virus long terminal repeat were generated and cross-bred with animals that highly express a bovine α -lac transgene (0.4 mg of α -lac· ml-1 of milk). The ribozyme contains the hammerhead catalytic domain, flanked by 12-nt sequences complementary to the 3′ untranslated region of bovine α -lac transcript. High-level expression of the ribozyme gene was detected by Northern blot analysis in the mammary gland of 7-8 day lactating transgenic mice, from 3 of 12 lines analyzed. Heterozygous expression of the ribozyme resulted in a reduction in the levels of the target mRNA to 78, 58, and 50% of that observed in the nonribozyme transgenic littermate controls for three independent lines. The ribozyme-mediated reduction in the levels of the bovine protein paralleled that observed for the mRNA, and was positively correlated with the level of expression of the ribozyme. In nonribozyme expressing transgenic mice, the level of bovine α -lac mRNA and protein was not affected. The specificity of this activity is demonstrated by the absence of a reduction in the levels of the endogenous murine α -lac mRNA or protein. These results demonstrate the feasibility of ribozyme-mediated down-regulation of highly-expressed transcripts in transgenic animals.