Department of Molecular Genetics and Microbiology
Animals; B-Lymphocytes; Base Sequence; Cell Line; DNA; DNA Primers; Escherichia coli; Immunoglobulin Isotypes; *Immunoglobulin Switch Region; Mice; Mice, Inbred BALB C; Mice, Nude; Molecular Sequence Data; Plasmids; Polymerase Chain Reaction; Transcription, Genetic; Transfection; Transformation, Genetic
Life Sciences | Medicine and Health Sciences | Women's Studies
Immunoglobulin class switch recombination (SR) occurs by a B cell-specific, intrachromosomal deletional process between switch regions. We have developed a plasmid-based transient transfection assay for SR to test for the presence of transacting switch activities. The plasmids are novel in that they lack a eukaryotic origin of DNA replication. The recombination activity of these switch substrates is restricted to a subset of B cell lines that support isotype switching on their endogenous loci and to mitogen-activated normal splenic B cells. The factors required for extrachromosomal plasmid recombination are constitutively expressed in proliferating splenic B cells and in B cell lines capable of inducibly undergoing immunoglobulin SR on their chromosomal genes. These studies suggest that mitogens that induce switching on the chromosome induce accessibility rather than switch recombinase activity. Finally, we provide evidence for two distinct switching activities which independently mediate mu-->alpha and mu-->gamma3 SR.
immunoglobulin, isotype switch, plasmid assay, transient transfection, PCR
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DOI of Published Version
J Exp Med. 2000 Apr 17;191(8):1365-80.
The Journal of experimental medicine
Shanmugam, A.; Shi, M. J.; Yauch, L.; Stavnezer, Janet; and Kenter, A. L., "Evidence for class-specific factors in immunoglobulin isotype switching" (2000). Women’s Health Research Faculty Publications. 191.
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