Question

1. You are conducting some in vitro splicing reactions with three different RNAs (A, B, and C) that each have a single intron. The exons in each case are identical in size (although the sequences could be different). One RNA has a group I intron, another has a group II intron, and the third has a spliceosomal intron. You incubate each RNA in conditions that allow self-splicing [(-) lanes below] as well as with a nuclear extract/splicing extract. After incubation, you observe the products by denaturing PAGE and autoradiography (your RNAs were radioloabeled). Which RNA has which intron

Answer 1

Step-by-step explanation:

1. There are six lanes in total out of which three are being showing self-splicing activity and the remaining three are being provided with splicing enzymes. As can be seen in case of RNA A, in the absence of a nuclear extract or when no splicing enzymes are added, there would be no splicing which can be reflected in lane 1 showing formation of only a single band which indicates that splicing does not occur on its own, although when the nuclear extract containing the splicing enzyme, there will be separation of the exons and the introns leading to the formation of two bands. So, the first two lanes specific for RNA A will be having a Spliceosomal Intron.

2. Now the task remains to identify which out of the remaining RNA's will be either Group I or Group II introns. There is no data regarding the addition of Guanosine nucleotide which is usually required in case of Group 1 splicing. Now, as we know that the splicing mechanism in case of Group II and spliceosomal intron is similar being carried out in a way in which an Adenine which is located in the branch site shows binding to the 5'- splice site which shows a similar conformational change for both the groups of introns. So, the Ribosome 2 which will act as a ribozyme and specifies lane 3 and 4 should be Group II intron and thus the last two lanes namely 5 and 6 will specify Group I intron.