Analysis of the Human T Cell Receptor α/δ Locus: New Approaches to Mapping and Sequencing

Citation

Boysen, Cecilie (1996) Analysis of the Human T Cell Receptor α/δ Locus: New Approaches to Mapping and Sequencing. Dissertation (Ph.D.), California Institute of Technology. doi:10.7907/7x0h-qy95. https://resolver.caltech.edu/CaltechTHESIS:12162020-194820334

Abstract

The human T cell receptor (TCR) α/δ locus has been mapped and sequenced. This region occupies roughly one megabase (Mb) of DNA or equivalent to one three thousandth of the entire human genome, the longest continuous piece of human DNA yet sequenced. The sequence has provided new insights into the complex organization, structure and evolution of two intermingled multigene families (α and δ), and will hopefully in the future help answer interesting questions concerning the complex expression patterns of TCR α and δ chains and about possible associations between specific polymorphisms in the TCR α/δ locus and susceptibility to autoimmune diseases. Comparison to cDNA data has provided information about expression of each of the TCR elements and about the striking diversification in the third hypervariable or junctional region. The sequence has contributed a glimpse of closely associated genomic DNA, in that the sequences surrounding the TCR locus, include the defender against death gene as well as five olfactory receptor genes. The sequence also harbors many other stretches of DNA, highly similar to previously identified genes, although in most cases, these have been found to be nonfunctional due to one or a few mutations. Comparison of 130 kilobases (kb) in the 3' region of the human sequence with its murine counterpart, suggests this region is highly conserved. The same 3' region has also been found to be limited in the concentration of genome wide repeats compared to the remainder of the locus. Furthermore, it contains a substantially reduced frequency of DNA variations compared to the rest of the locus. Apart from DNA variations in noncoding sequence, polymorphisms have also been identified in the coding regions of the TCR variable (V) gene segments, where, if they lead to amino acid changes, may alter the function of the TCR.

During the physical clone mapping and sequencing, new strategies were tested using primarily bacterial artificial chromosome (BAC) clones. These clones proved to be much more reliable and stable than clones currently employed in the human genome project (e.g., cosmids and yeast artificial chromosomes, YACs). BAC inserts can be sequenced completely by the high redundancy shotgun approach. Their insert size, stability, and capacity to be easily sequenced suggests that BAC clones are excellent mapping and sequencing reagents. The ends of BAC clone inserts can be sequenced directly. This has led to the proposal of a new strategy for obtaining the entire DNA sequence of the human genome without physical mapping.

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