Team:Penn State/Bidirectional Promoters Design
From 2012.igem.org
Presentation
Scientists are frequently confounded by wayward promoters; that is, promoters which do not produce the expected proteins. Some bidirectional promoters are known to exist, but which way they promote and the degree of expression has not been quantified. This project will test the directionality of several BioBrick promoters to answer these questions.
Bidirectional Promoters
Background
All of the proteins around us, with few exceptions, are made up of 20 fundamental building blocks of life - amino acids. Different arrangements and combinations of these basic building blocks give us the diversity of proteins that we see. Messenger RiboNucleic Acids (mRNA) is in essence a "photocopy" of DNA that codes for a gene. mRNA carry codons, which are groups of three bases that code for a single amino acid. There are 64 possible combinations of codons (4 x 4 x 4 = 64), but these combinations are degenerate, so there can be more than one codon that codes for a single amino acid.
The Problem
Proteins are assembled by ribosomes which read the mRNA and catalyze the binding between Amino Acids; however, they cannot read the codon code themselves and require the help of tRNA (transfer RNA). Each tRNA matches a single codon sequence and can only carry a single specific amino acid at a time. tRNA has complimentary bases to their respective codon sequences on the mRNA strand, and once the tRNA matches the sequence on the mRNA, the tRNA deposits the Amino Acids, which then bind together to form proteins.
We have mentioned how these can be a number of codons that can code for the same Amino Acid, and subsequently, a number of tRNA molecules that can carry a given Amino Acid. In nature, and in many organisms, only a select few of these tRNA and codon combinations are used instead of all sequences that code for the same amino acid. This is called codon bias.
Objective
Our goal for this project is to see which codons are preferred, or biased. We also want to investigate if this bias can change over time, different circumstances, or stresses.