Question

A1. The following is the DNA sequence of a hypothetical gene for the SMALL protein. It is called the SMALL gene. 1 atgggattac actgtcacga ccaaatagcc ttcattgtat 41 caaaaggatc aatcgagtta tag Imagine you are doing a research project in a laboratory and your supervisor asks you to clone the SMALL gene into the pBR322 plasmid (shown below) You must use the Pstl and EcoRI sites for your cloning HindIII EcORIECORV BamHI 359 0 29 185 4000 375 Sall PstI 651 3607 tet amp 1000 pBR322 4361 bp 3000 ori 2000 2295 Ndel Your first step in the cloning is to cut pBR322 with Pstl and EcoRI (in the same tube) then electrophorese the cutting mixture on an agarose gel to examine the cutting efficiency. A picture of the gel electrophoresis equipment is shown on the next page.

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Answer 1

Answer.a] electrode A is negatively charged because DNA is loaded in well at A electrode. As we know that DNA has phosphate group which make it negatively charged. So DNA will move towards positively charged electrode.

Answer b] Reverse primer = 5' CTA TAA CTC GAT TGA CC 3' for making reverse primer we used the 3' end of the DNA and reverse the sequence in this at 3' the sequence is TAG and reverse of it will be 5' CTA, so as such we can make reverse primer.

Answer c] PCR use thermostable DNA polymerase because PCR cycle runs at higher temperature which is 72 Degree celcius. While denaturation of DNA occurs at 92 degree celcius. So a thermostable DNA polymerase is needed so that DNA polymerase will not denatured at high temperature.

Answer d] there are 63 bases and out of which one is stop codon so total bases remains are 60 also we know 3 bases codes for one amino acid so amino acid residue encoded in protein 60/3 = 20 amino acid longs.