Rapid cloning of any rearranged mouse immunoglobulin variable genes

Journal Article · Tue Dec 31 04:00:00 EST 1996 · Immunogenetics

DOI:https://doi.org/10.1007/BF00176675· OSTI ID:501870

Dattamajumdar, A K ^[1]; Jacobson, D P; Hood, L E; Osman, G E ^[2]

---

1. Univ. of Washington, Seattle, WA (United States)
2. Univ. of Washington School of Medicine, Seattle, WA (United States)

Immunoglobulins (Ig) have been the focus of extensive study for several decades and have become an important research area for immunologists and molecular biologists. The use of polymerase chain reaction (PCR) technology has accelerated the cloning, sequencing, and characterization of genes of the immune system. However, cloning and sequencing the Ig variable (V) genes using the PCR technology has been a challenging task, primarily due to the very diverse nature of Ig V region genes. We have developed a simple, rapid, and reproducible PCR-based technique to clone any rearranged mouse Ig heavy or light chain genes. A close examination of all Ig heavy and light chain V gene families has resulted in the design of 5{prime} and 3{prime} universal primers from regions that are highly conserved across all heavy or light chain V gene families, and the joining or constant regions, respectively. We present our strategy for designing universal primers for Ig V gene families. These primers were able to rapidly amplify the rearranged Ig V genes, belonging to diverse Ig V gene families from very different cell lines, i.e., J558, MOPC-21, 36-60, and a chicken ovalbumin specific B-cell hybridoma. In addition, the present study provides the complete alignment of nucleotide sequences of all heavy and light chain variable gene families. This powerful method of cloning Ig V genes, therefore, allows rapid and precise analysis of B-cell hybridomas, B-cell repertoire, and B-cell ontogeny. 55 refs., 5 figs., 2 tabs.

• Sponsoring Organization: USDOE
• OSTI ID: 501870
• Journal Information: Immunogenetics, Journal Name: Immunogenetics Journal Issue: 3 Vol. 43; ISSN 0093-7711; ISSN IMNGBK
• Country of Publication: United States
• Language: English

Related Subjects

44 INSTRUMENTATION
INCLUDING NUCLEAR AND PARTICLE DETECTORS
55 BIOLOGY AND MEDICINE
BASIC STUDIES
AMINO ACID SEQUENCE
ANTIGENS
DNA SEQUENCING
DNA-CLONING
EFFICIENCY
EVALUATION
GENE MUTATIONS
GENETIC MAPPING
HEREDITARY DISEASES
HYBRIDOMAS
IMMUNE REACTIONS
IMMUNOGLOBULINS
LYMPHOCYTES
MICE
NUCLEOTIDES
POLYMERASE CHAIN REACTION
QUANTITATIVE CHEMICAL ANALYSIS
RECEPTORS
RELIABILITY
STRUCTURE-ACTIVITY RELATIONSHIPS