Question

You are doing an experiment in which you are using an in vitro translation system derived from rabbit reticulocytes, called a rabbit reticulocyte lysate, to translate mRNA in a test tube. The rabbit reticulocyte lysate contains ribosomes, translation initiation and elongation factors, tRNAs, aminoacyl tRNA synthetases, release factors, amino acids, and ATP. The lysate does not contain any mRNA, so you must add the mRNA you would like to translate to the system. A. When you analyze the proteins made from your lysate, you find that they are all only two amino acids long. At first, you think that there must be a mutation to a stop codon in the position of the third codon, so you do another experiment with a different mRNA. Again, all the newly synthesized proteins are only two amino acids long, so you conclude that there is a problem with some component in your lysate.
Describe ONE possible problem in the translation system (lysate) that would result in the limitation of newly synthesized peptides to two amino acids in length.

Answer 1

Answer:

Problems with transfection efficiency or absence of essential mRNA modifications (capping and poly-A tail)

Explanation:

Translation is the process of protein synthesis from RNA in which the genetic information encoded in a messenger RNA (mRNA) molecule is translated into a protein sequence composed of amino acids. Translation has three steps: initiation, elongation, and termination. During translation, different elements are required to initiate, continue and complete the process, which include essential molecules such as amino acids, mRNA, tRNAs, ribosomes, energy-carrying molecules (i.e., ATP, GTP), initiation factors (i.e., eIF1A, eIF3, etc) and elongation factors (eEF-2, EIF5A). For example, humans can't synthesize nine essential amino acids (i.e., threonine, tryptophan, phenylalanine, valine, methionine, leucine, histidine, isoleucine, lysine), thereby it is imperative to include these amino acids to synthesize proteins composed of them. On the other hand, it is also fundamental to take into account that the efficiency of transfection of the molecules required during translation, as well as the absence of modifications in the mature mRNA sequence, may alter the process in vitro.