Targeting expression to the mammary gland: intronic sequences can enhance the efficiency of gene expression in transgenic mice |
| |
Authors: | C Bruce A Whitelaw Alan L Archibald Stephen Harris Margaret McClenaghan J Paul Simons A John Clark |
| |
Institution: | (1) Present address: Edinburgh Research Station, AFRC Institute of Animal Physiology and Genetics, EH25 9PS Roslin, Midlothian, Scotland, UK |
| |
Abstract: | We are studying the tissue-specific expression of the sheep milk-whey protein gene, β-lactoglobulin. We have used sequences
derived from this gene to target the expression of biomedical proteins into milk with the intention to exploit this technology
in transgenic sheep as a means of protein production. In the present study, a series of β-lactoglobulin hybrid genes and β-lactoglobulin
minigenes were evaluated for expression in the mammary gland of transgenic mice. In particular, we have assessed whether there
is a requirement for introns for efficient transgene expression in the mammary gland, since the coding sequences of many candidate
proteins are available only as cDNAs. The results suggest that the inclusion of natural introns in constructs can enhance
the efficiency of transgene expression. Thus, a hybrid construct comprising 4.3 kb of the immediate 5′ flanking sequences
of β-lactoglobulin fused to a genomic minigene encoding human α-antitrypsin (α1AT) was expressed much more efficiently than an α1AT-cDNA construct containing the same β-lactoglobulin segment. Similarly, the intact β-lactoglobulin gene was expressed more
efficiently than the corresponding intronless β-lactoglobulin minigene. This effect was not seen in transient expression expriments
in baby hamster kidney cells when β-lactoglobulin-α1AT constructs were driven by SV40 enhancer sequences. The effect cannot be explained by a simple requirement for splicing,
since the inclusion of the first β-lactoglobulin intron into cDNA constructs encoding human α1AT or β-lactoglobulin itself failed to enhance the efficiency of transgene expression. It is concluded that sequence elements
within introns may interact with the upstream 5′ flanking sequences of β-lactoglobulin and enable the latter to function efficiently
in the mammary gland of transgenic mice. |
| |
Keywords: | β -lactoglobulin tissue specificity hybrid genes RNA splicing intervening sequences |
本文献已被 SpringerLink 等数据库收录! |
|