Genetically manipulated mice: a powerful tool with unsuspected caveats

Abstract

Although genetic manipulations in mice have provided a powerful tool for investigating gene function in vivo, recent studies have uncovered a number of developmental phenomena that complicate the attribution of phenotype to the specific genetic change. A more realistic approach has been to modulate gene expression and function in a temporal and tissue-specific manner. The most common of these methods, the CreLoxP and tetracycline response systems, are surveyed here and their recently identified shortcomings discussed, along with a less well known system based on the E. coli lac operon and modified for use in mammals. The potential for further complications in interpretation due to hitherto unexpected epigenetic effects involving transfer of RNA or protein in oocytes or sperm is also explored. Given these problems we reiterate the necessity for the use of completely reversible methods that will allow each experimental group of animals to act as their own control. Using these methods with a number of specific modifications to eliminate non-specific effects from random insertion sites and inducer molecules, the full potential of genetic manipulation studies should be realized.

Growth Arrest Specific 1 (GAS1) Is Abundantly Expressed in the Adult Mouse Central Nervous System

“…In this case, the transgenic mouse expresses a reporter gene (GFP) under the control of a UTR DNA sequence 5` of the GAS1 coding region, contained in a BAC. Whereas this type of mouse has greatly reduced the positional effect of transgene insertion, it is not yet clear how the presence of other genes, transcription factors, microRNAs, or controlling regions present in these large DNA fragments may alter the expression of the protein under study (Matthaei 2007). Furthermore, the processing of the reporter protein is different from that of the endogenous protein: in this case, the reporter protein is soluble, thus its intracellular distribution is distinct from the endogenous (GAS1) protein.…”

Unsuspected effects of a lung‐specific cre deleter mouse line

“…This second Sftpc-cre mouse strain was established in the ICR genetic background. The difference in behavior of the two transgenic lines can either be due to variation in the genetic background, or more likely, to distinct random integration event of the transgene in the genome that can affect transgene expression levels (Matthaei, 2007;Mura et al, 2010;Okudo et al, 2005).…”

“…Over the last decade, the number of Cre-expressing mouse lines has been growing steadily allowing the research community to have access to an important resource with a strong impact on our comprehension of mammalian genetics. However, unanticipated caveats observed with the use of Cre-expressing mouse lines demonstrate the limits of the approach and how crucial experimental controls are (Koitabashi et al, 2009;Matthaei, 2007;Naiche and Papaioannou, 2007;Schmidt et al, 2000;Whitsett and Perl, 2006).…”

Use of Tissue Cross-reactivity Studies in the Development of Antibody-based Biopharmaceuticals

“…Overall, it is not recommended to conduct TCR studies with test articles that do not contain a CDR. Furthermore, although IHC may be used to justify research use of a genetically engineered model, full tissue panel TCR studies in these animals are not recommended because protein distribution and expression in these models are influenced by the design of the transgene and the gene integration site(s), and they may not reflect the epitope as expressed in the human or the selected toxicology species (Matthaei 2007).…”