Essential genes in a region of chromosome I in Caenorhabditis elegans

• Main Author: Howell, Ann Marie
• Language: English
• Published: University of British Columbia 2010
• Online Access: http://hdl.handle.net/2429/29113

This thesis describes the identification and characterization of essential genes in a small region of the genome of the nematode Caenorhabditis elegans. The region analyzed was defined by a 1.2 map unit deficiency of chromosome l, hDf6. In order to do this, a system for recovering and analyzing a large number of autosomal recessive lethal mutations was developed. This system used a free duplication of the left third of chromosome I, sDp2. Lethal mutations were maintained with two mutant alleles on the normal chromosomes and a wild-type allele on the duplication. More than 31,000 chromosomes mutagenized with ethyl methane sulfonate were screened using the sDp2 system and 495 lethal mutations were recovered. Two translocations involving chromosome I (hT1(l;V) and szT1(l;X), were also used to recover lethal mutations. It was discovered that heterozygotes for one of these translocations, szT1(l;X), exhibited an increased frequency of recombination adjacent to the chromosome I breakpoint. This had not been observed previously with other translocations. The increase in recombination frequency may be the result of a disruption of the normal regulation of recombination in the region. Recombination mapping was used to position 59 EMS-induced sDp2-recovered lethal mutations. Deficiency mapping, duplication mapping, and inter se complementation analyses were used to define 36 essential genes to the left of the dpy-5 gene on chromosome I. Nineteen of these genes, defined by 54 lethal mutations, were uncovered by the deficiency hDf6. A small gamma radiation-induced deficiency, hDf7, was identified and found to uncover six of the essential genes in hDf6. Two duplications which have breakpoints in the hDf6 region, hDp3 and hDp25, were also used to position essential genes. The genes in hDf6 were thus positioned into five regions based on the breakpoints of these three other chromosomal rearrangements. The stage of lethal arrest was determined for the lethal mutations in hDf6. Mutations in genes in the left portion of hDf6 arrest development at an earlier stage than do mutations in genes in the right portion. As a whole, the hDf6 region seems to have a lower proportion of early arresting mutations than other comparably sized regions of the genome. Ten of the 19 genes in hDf6 were represented by more than one lethal allele. One gene, let-354, was found to be an extremely mutable target since it was represented by seventeen alleles. A truncated Poisson analysis of the allele distribution indicated that the 19 genes identified represented a maximum of 75% of the essential genes predicted to be in hDf6. Extrapolating from this region to the entire genome, an estimate of 4,000 essential genes was obtained. The average forward mutation rate was determined to be 5 x 10⁻⁵ mutations per gene. It was estimated that 60,000 chromosomes mutagenized with 0.015 M EMS must be screened to identify mutationally most of the genes in any region of the genome. Thus, it would require the screening of another 30,000 mutagenized chromosomes to identify genetically almost every gene in the hDf6 region of C. elegans. === Medicine, Faculty of === Medical Genetics, Department of === Graduate