 So far we have discussed that DNA carries the genetic information in the form of nucleotide sequence. How is this information, genetic information communicated to rest of the cellular machine? How genetic information influences the functioning of a cell? A CD, a compact disc can contain several songs but the songs will not play on their own. DNA requires specific machinery to do that. Similarly DNA also requires certain machinery to be able to communicate its information to the cell and modify or change the cellular function. So happens that people realized that the molecular basis of phenotypes were discovered before it was known that DNA is the genetic material. We will start with the experiments of Archie Paul Gerard. In 1908 he was studying a disease called Alcaptanuria or Black urine disease. He noticed that there was precedence of children of consignuous marriage between two cousins or close relatives. These children had a higher rate of suffering from this particular disorder. So he collected samples, urine samples and blood samples and discovered that these people were missing a particular enzyme, a protein. The name of that protein is homogenistic acid oxidase. It converts tyrosine into homogenistic acid and in metabolism, tyrosine is converted into homogenistic acid which is then converted into harmless product. So this acid basically neutralizes homogenistic acid. Though levels of homogenistic acid can be tolerated but if this enzyme homogenistic acid oxidase is missing or non-functional homogenistic acid will accumulate and it will cause damage. So this gentleman was able to conclude and he was able to link biochemical phenotype of a disease to an abnormal gene and a missing enzyme which was missing in the blood samples. Now let's move on a little bit in time. 1940s, the classic experiment of Betel and Tatum. These gentlemen were studying a specific type of mold called neurospora. They basically did experiments, they took this organism, this mold and they mutated this mold using X-rays. X-rays were used as a mutagen and that resulted in an altered phenotype and they were able to associate altered phenotype with an altered enzyme or enzyme activity. So generally neurospora can grow on minimal media, that is minimal amount of amino acids, carbohydrates, lipids, etc. required for this organism to grow. When they used X-rays to mutate this organism, some of these mutated neurospora strains they were unable to grow on minimal media, they had to supplement it. So the term prototrophs, the original eaters or oxotrophs, the increased eaters, they came up with those terms. So if they realized that if they supplemented the media with certain amino acids or certain chemical compounds, they were able to grow those mutants. Let's look at that. So mutants that were arginine, which required arginine plus the minimal media to grow, they called them arch mutants and they saw the different versions of arch mutant present in the specimen. So they realized that if arginine is the compound that is missing, let's see if the precursor to arginine can also recover these mutants. So they classified each mutation as affecting one enzyme or another in the metabolic pathway that results in production of arginine in the cell. Wild type, meaning the original strain which did not require any special nutrients to grow and the mutant cells, they were examined for enzyme and their activities using biochemical assays. The results confirmed that each mutant strain was indeed missing a single active enzyme in the pathway of arginine production. Here's a photograph of their experiment. You can see there are the wild type, it can grow in minimal media in which no supplements, additional supplements are added. So the wild type can grow in even if the additional supplements are added. There's strain number one or mutant strain number one. It could not grow unless and until arginine was provided. Even the products ornithine and citrulline which are intermediates in arginine biosynthetic pathway, they could not recover these mutants. Other arginine mutants could be recovered not only by arginine but with also citrulline. These mutants could grow if citrulline or arginine either one of them was provided but could not grow if they were just given ornithine they could not grow. The third mutant strain needed ornithine wasn't provided, they were not able to grow and citrulline and arginine they also if that was these two compounds are also provided they could grow in these compounds also. So here is the interpretation. If an organism cannot convert a particular compound to another it presumably lacks an enzyme required for the conversion and the mutation is the gene that codes for that enzyme. Here you can see there is a precursor molecule which is converted first into ornithine by an enzyme A coded by gene A. Ornithine is then converted into citrulline by enzyme B and then also citrulline is ultimately converted into arginine by enzyme C. So the interpretation is that our if the strain had mutation somewhere here it could not convert ornithine into citrulline then we would need either citrulline or arginine to grow these mutant strains. So rest is simple and straightforward. So this experiment was one of the initial experiments that linked change in gene to a function or ability of cells to grow and connecting the phenotype with the genotype through an enzyme.