Q & A: Isolation of Genomic DNA

Q1: Why Agarose is used instead of Agar?

Ans: Agarose is polymer of D galactose and 3, 6 anhydro 1 galactose. Agar is polymer of agaro pectins (Pore size will small).

Q2: How alcohol is precipitate DNA?

Ans:  Dehydration.

Q3: Why 70 % ethanol is used for cleaning? Why no 60%, 80%, 90%?

Ans:  30% of water dissolve the impurities.

Q4: What is the role of BPB?

Ans:  BPB – Bromo Phenol Blue. Tracking the DNA

Q5: What the components of gel loading dye?

Ans: Tris (pH - 8), BPB and sucrose

Q6: Why TBE buffer?

Ans: Tris borate to maintain the pH (8.6); EDTA protect DNA.

Q7: why TE buffer is pH - 8?

Ans: Both DNA & RNA are acid. To store them, by inactivating catalyses.

Q8: Why we use Proteinase K? what K stands for?

Ans: To degrade the protein. ‘K’ - keratin

Q9: Why we are using RNase A and not RNase H?

Ans: RNase use to cleave the RNA. RNase H is used cleave double stand RNAs.

Q10: What is the source of enzyme we used today?

Ans: Lysozyme/Proteinase K

Q11: why incubation at 37°C?

Ans: Most of the enzymes are highly active.

Q12: What happen if

  50x TBE used – high current & temp denature 

  water is used – no current will pass through

Q13: Why gel for gDNA analysis in less than 1%?

Ans: gDNA is large so it needs large pore size.

Q14: Why we isolate the DNA?

Ans: For sequencing, mutation and recombination etc

Q15: How Ethidium Bromide works?

Ans: It bind to the DNA double stand. In RNA makes the secondary structure (homodimer) and EtBr binds to that