TrackingH-2 alleles in transgenic mice by RFLP and heteroduplex analysis

Abstract

Transgenic mice provide valuable tools for biological research including many areas of immunology. In studies involving the major histocompatibility complex (MHC), it is often necessary to place the desired transgene in a specific H-2 (the murine MHC) environment. In this regard, the strains commonly used for the production of transgenic mice also carry well characterized H-2 alleles and provide an appropriate genetic background for MHC related experiments. In this study, a highly polymorphic microsatellite of tetranucleotide repeats from the second intron of the class II Eb gene within the H-2 complex was used in order to identify the corresponding alleles. The relevant H-2 allele(s) along with the transgene were then tracked throughout the production of a chicken ovalbumin-specific transgenic strain. The technique involved PCR-amplification of a DNA sequence encompassing the H-2 specific microsatellite followed by RFLP and heteroduplex analyses. This approach is likely to find wide application in the background checking of transgenic mice, especially in immunological research requiring a defined H-2 background.

Tiermodelle in der Transplantationsimmunologie: Screening auf MHC-Antigene hoher Homologie durch RFLP-Analyse als Alternative zur Durchflußzytometrie

“…Damit bietet sie einen entscheidenden Vorteil zu der von Shanmugam et al vorgestellten Methode eines Restriktionsdigest fUr H-2 Allele, welche durch die minimalen Langenunterschiede der Schnittfragmente keine eindeutige Aussage bei heterozygoten Tieren erlaubt [5]. Bei vergleichbarem Zeitaufwand und geringeren Kosten erwies sich dagegen die RFLP-Analyse -bei Vorliegen entsprechender Restriktions-Polymorphismen -als zum Screening nahezu homologer MHC Genotypen besonders geeignet.…”