Tafaani Khan Section 009
March 26, 2018 Manuscript II
Our hypothesis in question was to test the PTC genotype and determined PTC phenotype of individuals in a large population. Our analysis led to an understanding of whether the determined genotypes accurately related to the expected PTC phenotypes. We tested for Hardy-Weinberg equilibrium. We performed PCR (to amplify the specific PTC locus) and a restriction digest that would cut the taster but not the non-taster allele (i.e. identify the polymorphism). The restricted PCR reaction, of each individual, was loaded onto an agarose gel. The fragments of each sample separated based on size, where the dye aided in visualization and the molecular weight markers served as references. We used multiple paper strips to differentiate between tasters/non tasters. If the PTC paper was bitter, then we were tasters, in contrast to those who were nontasters (no bitter taste evident).
We show that individuals in the population were homozygous tasters (Fig. 1, Lanes 5 and 7), homozygous non-tasters (Fig. 1, Lanes 3, 8, and 9), and heterozygous (Fig. 1, Lanes 1, 2, and 4). Overall, we saw correlation between PTC genotype and expected phenotype, with two non-correlation events. One individual was qq (non-taster genotype) with a taster phenotype (Table 1: Genotype determines phenotype). Another individual was pq (taster genotype) with a non-taster phenotype (Table 1: Genotype determines phenotype). We employed the Hardy-Weinberg equation to conclude if our population was under selective evolutionary pressure. The population is not in HWE (the population is evolving so the allele frequencies are changing). The observed genotypic frequencies, p= 0.61 and q= 0.39, deviate from the expected phenotypic frequencies, p= 0.64 and q= 0.36 (Table 1: Genotype determines phenotype). Thus, the frequency of p and q based on phenotypic analysis of the PTC allele does not correlate with the frequency of each based on molecular data. Assuming a random population follows Hardy Weinberg Equilibrium, allele frequency would be 50/50, which is not evident.
The determined genotypes corresponded to the expected phenotypes, except for two instances where determined genotype did not correspond to the phenotype. The overall population was not in Hardy Weinberg Equilibrium, as the allelic frequencies were not 50/50, as expected for a random mating population. As in the article, “Independent evolution of bitter-taste sensitivity in humans and chimpanzees”, chimpanzees and humans both possess apparent dominant and recessive ‘taster’ and ‘non-taster’ alleles at roughly equivalent frequencies. However, a “maximum-likelihood phylogeny revealed that TAS2R38 haplotypes in humans and chimpanzees form separate clades” (non-taster allele in each species is derived from an ancestral taster allele). This concept of haplotypes would explain the unique cases of the two individuals in our population. In “Positional Cloning of the Human Quantitative Trait Locus Underlying Taste Sensitivity to Phenylthiocarbamide”, haplotypes accounted for the detected variance in PTC sensitivity, where amino acid substitutions at different positions affected the relationship between PTC genotype/phenotype. For the individual who was homozygous recessive (qq) but could taste PTC, a possibility for this phenomenon is that FnuH1 restriction enzyme was not added. For the individual who was heterozygous (pq) and could not taste PTC, a possibility for this phenomenon is due to haplotypes (where all 3 SNPs (PAV) were not inherited to detect PTC). Another possibility is the restriction enzyme digest worked but there was genetic contamination (more of someone else’s DNA than themselves, which is easier to disrupt the gene).
Substantial variation in taste sensitivity exists in humans, where phenotypic variation has an underlying genetic origin. Taster and non-taster alleles have been persistent in human populations, as detecting bitter substances is beneficial (identifying poisonous substances) just as not detecting bitter substances is beneficial (consuming nutritious foods). While the experiment ran successfully, it is important to use caution when adding the restriction enzymes, inserting the restricted PCR reaction in the gel lanes, and preventing genetic contamination. The haplotypes explain the bimodial distribution of PTC taste sensitivity, accounting for differing abilities to taste PTC (some more strongly than others). Overall, genetics is the driving force behind expected phenotypic expression; modifying genomic sequences provides great insight for the concept of forward genetics. Presently, we know that the ability to taste PTC (or not) is presented by a single gene that codes for a taste receptor on the tongue (TAS2R38) (i.e. bitter taste receptors are more concentrated on the sides of your tongue). Most importantly, the interesting nature of PTC and the study of gene-trait relatedness provided new understandings towards the central dogma of molecular biology.
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