Genomic complexity of the variable region-containing chitin-binding proteins in amphioxus

<p>Abstract</p> <p>Background</p> <p>The variable region-containing chitin-binding proteins (VCBPs) are found in protochordates and consist of two tandem immunoglobulin variable (V)-type domains and a chitin-binding domain. We previously have shown that these polymorphic genes, which primarily are expressed in the gut, exhibit characteristics of immune genes. In this report, we describe VCBP genomic organization and characterize adjacent and intervening genetic features which may influence both their polymorphism and complex transcriptional repertoire.</p> <p>Results</p> <p>VCBP genes 1, 2, 4, and 5 are encoded in a single contiguous gene-rich chromosomal region and VCBP3 is encoded in a separate locus. The VCBPs exhibit extensive haplotype variation, including copy number variation (CNV), indel polymorphism and a markedly elevated variation in repeat type and density. In at least one haplotype, inverted repeats occur more frequently than elsewhere in the genome. Multi-animal cDNA screening, as well as transcriptional profilingusing a novel transfection system, suggests that haplotype-specific transcriptional variants may contribute to VCBP genetic diversity.</p> <p>Conclusion</p> <p>The availability of the <it>Branchiostoma floridae </it>genome (Joint Genome Institute, Brafl1), along with BAC and PAC screening and sequencing described here, reveal that the relatively limited number of VCBP genes present in the amphioxus genome exhibit exceptionally high haplotype variation. These VCBP haplotypes contribute a diverse pool of allelic variants, which includes gene copy number variation, pseudogenes, and other polymorphisms, while contributing secondary effects on gene transcription as well.</p

Vector system for selection of genes encoding secreted proteins and membrane-bound proteins

The subject invention concerns novel vectors for the rapid and robust selection for cDNA sequences that encode secreted or membrane-bound proteins. The invention also pertains to methods for cloning secreted or membrane-bound proteins, including proteins encoded by novel members of gene families

Characterization of three isotypes of immunoglobulin light chains and T-cell antigen receptor α in zebrafish

The zebrafish (Danio rerio) has become a significant model for understanding the developmental regulation of gene expression and holds considerable potential for characterizing the development of the immune system. Using a number of different approaches, including heterologous hybridization and short-primer PCR, cDNAs for three different classes of light-chain genes were identified and characterized. The zebrafish light chains are similar to trout type 1, trout type 2, and catfish type F, respectively. T-cell antigen receptor α (TCRα) was also identified and characterized. A high proportion of unusual transcripts including sterile transcripts, germline VJC transcripts, aberrant splice forms, and V-V transcripts were encountered in the immunoglobulin and TCR cDNAs examined. The light-chain and TCRα loci each consist of multiple families of V gene segments, apparent even from the small numbers of cDNAs of each isotype sequenced. The gene sequences reported provide an essential set of markers of both B- and T-cell lineages that will facilitate investigations of immune system development

Characterization of three isotypes of immunoglobulin light chains and T-cell antigen receptor α in zebrafish

The zebrafish (Danio rerio) has become a significant model for understanding the developmental regulation of gene expression and holds considerable potential for characterizing the development of the immune system. Using a number of different approaches, including heterologous hybridization and short-primer PCR, cDNAs for three different classes of light-chain genes were identified and characterized. The zebrafish light chains are similar to trout type 1, trout type 2, and catfish type F, respectively. T-cell antigen receptor α (TCRα) was also identified and characterized. A high proportion of unusual transcripts including sterile transcripts, germline VJC transcripts, aberrant splice forms, and V-V transcripts were encountered in the immunoglobulin and TCR cDNAs examined. The light-chain and TCRα loci each consist of multiple families of V gene segments, apparent even from the small numbers of cDNAs of each isotype sequenced. The gene sequences reported provide an essential set of markers of both B- and T-cell lineages that will facilitate investigations of immune system development

DNA vector for determining the presence of out-of-reading-frame mutations

A DNA expression vector for positively selecting in-frame or out-of-reading-frame mutations in DNA sequences to be tested comprising a promotor operatively linked to an expressible reporter gene through a linkage sequence is disclosed. The linkage sequence includes at least two restriction sites and an engineered frameshift mutation. In an embodiment the frameshift is established by complementary sequences SEQ ID Nos:1 and 2. The expressible reporter gene is expressed as a fusion product including a green fluorescent protein and the promoter can be lacZ and inducible in E. coli