When using the genetic map the DNA sequence analysis in this experiment, they were able to line up the genetic maps of the maize and the rice and identify exons of maize that match with the rice. After this analysis the open reading frame shows that the amino acids in the maize is 58% identical and 66% similar to the rice. When they used another tgal allele for the same test they found that it confirmed that tgal is the SBP gene, and it shows that “a single amino acid substitution is sufficient to confirm the difference between the maize and teosinte phenotypes” (pg 715). These differences could be due to the tgal analysis or the gene expression. The next test was the northern blot analysis. This is another gene expression test. They compared this expression to the patterns in the in situ hybridization. There was a weak, but distinctive expression in this test. It showed that the immature ear of both the maize and the teosinte were different. The genetic analysis shows that they can compare the base pairs of the amino acids to prove or disprove that the maize and teosinte are related. When using the western blot they saw that the protein programmed in the teosinte was more abundant during the developmental stages of the plant. In the last test called selective sweep, the nucleotide diversity was compared as illustrated in graph 5a. A selective sweep is the reduction or elimination of variation among the nucleotides in neighboring DNA of a mutation as the result of recent and strong natural selection. It was found that the teosinte is similar to the maize, but the promoter maize is less diverse than the promoter in the teosinte. The ratio of diversity shows that the promoter region in the maize tbl gene, another maze domestication gene, possesses only 5% of the diversity found in teosinte. During the selective sweep the differences between the maize and teosinte are in the promoter and exon 1, but in exon 2, 3 and so on, the two plants are similar.
The purpose of this set of experiments was to determine what genetic differentiation is responsible for the change in a covered kernel to one that is exposed. In order to test this hypothesis, the genetic make-up of both plants were mapped and compared, gene assays were obtained and viewed, and nucleotide diversity between 16 land races of maize and 12 teosinte individuals were ascertained. These different methods of exploration led researchers to believe that a single substitution in the tgal protein-coding gene is responsible for this drastic phenotypic change.
It turned out that the factor(s) responsible for the change in phenotype between teosinte and maize happened to lie within a certain region of the plant’s genome. This region was responsible for the coding of seven DNA sequences which showed very little variation. Nonetheless the team of researchers found that one of the genes in the maize plant had been differed by one amino acid change than those of the teosinte. This change led them to believe that this had been the target they were looking for.
Once the parameters for testing their hypothesis had been set, the research team began to experiment. When trying to find the affects of this substitution on maize, the gene was reverted back to the original found in the progenitor. A clone of the subject (containing the reversal) was grown, and observations were taken on the cupules and glooms formed over the kernels. The results of this experiment have helped in the formation of a conclusion focusing on the idea that miniscule changes in the genotype of an organism may very well be responsible for dramatic changes in phenotype.
After narrowing down the causative site for functional differences between maize and teosinte to the 1,042 base pair segement, DNA sequence analysis was done on this segment. It identified seven fixed differences between maize and teosinte that could affect tga1 expression. The protein sequence which was determined through the protein assay method, allowed the pinpointing of the seventh difference which encodes an amino acid substitution of lysine (K) in teosinte to asparagine (N) in maize. This method helps assess the magnitude of phenotypic differences, by finding the region for which the single amino acid is different.
The research group responsible for this experiment believes that the gene they found was the one being selected on during the domestication of teosinte to a more commercial product.