Question

The diagram shows the size and position of the exons (numbered) and introns (lines) of a gene that codes for hypothetical Protein X, which can exist as two isoforms (either 16 or 17 amino acid residues long). Why " A" is the correct answer, can you please explain step by step. Thank you!

24 bp 9 bp 6 bp 18 bp

Which technique can be used to determine if a sample of cells expresses both isoforms?

1. Synthesize cDNA from Protein X mRNA using primers overlapping exons 1 and 4, followed by gel electrophoresis and band visualization.
2. Separate Protein X mRNA by gel electrophoresis and visualize band pattern using a DNA probe complementary to exon 3.
3. Perform a PCR of Protein X genomic DNA using primers overlapping exons 1 and 3, followed by gel electrophoresis and band visualization.
4. Perform a restriction digest of Protein X genomic DNA using an endonuclease that cuts in the middle of exon 2, followed by gel electrophoresis and band visualization.

Answer 1

Answer: Option A is correct
Explanation:
Total length of exons = 24+9+6+18
= 57 bp

Total number of codons in the mRNA = 19 = 18 sense codons + 1 termination codon
Different isoforms can be produced by alternative splicing.
The alternative splicing increases the repertoire of proteome and transcriptome without affecting the genome.

Protein with 16 amino acids is generated by skipping exon 2. (9 bp = 3 codons)
Protein with 17 amino acids is generated by skipping exon 3. (6 bp = 2 codons)

So, we can perform PCR using primers binding to regions 1 and 4 to determine the length of the matura transcript. For this purpose, we have to isolate mRNA from different tissues and synthesize cDNA.
If the primers anneal at the ends,
Isoform with 16 amino acids produces a band of 48 bp
Isoform with 17 amino acids produces a band of 51 bp