Which 3 of the following are true about DNA ligation? Choose at least one answer

Question

Which 3 of the following are true about DNA ligation?
Choose at least one answer.
a. Ligation of blunt ends is more efficient that ligation of sticky ends.
b. The invivo function of T4 DNA ligase is to repair single-strand nicks.
c. It requires energy in the form of dNTP.
d. It requires EDTA
e. A 5’ terminal phosphate is required.
f. It requires no input of energy. Energy is released by the joining of the sugar-phosphate backbone.
g. It requires energy in the form of ATP
h. A 3’ terminal phosphate is required.
i. T4 DNA ligase is most efficient at 20°C invivo.

Explanation / Answer

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RIGHT:
b. The in vivo function of T4 DNA ligase is to repair single-strand nicks.
e. A 5’ terminal phosphate is required.
g. It requires energy in the form of ATP

WRONG:
a. Ligation of blunt ends is more efficient that ligation of sticky ends. Nope! Sticky ends bind together much better than hard-to-ligate blunt ends. In fact, the longer the 'sticky end', the more efficient the ligation.

c. It requires energy in the form of dNTP. No. Neither dATP, dTTP, dGTP and dCTP are not used by the ligase. It is the ribose form ATP that is used. ATP is the common currency of energy in cells.

d. It requires EDTA. Nope. EDTA chelates divalent metal ions like Mg2+ and Ca2+. But the removal of these metal ions is neither beneficial nor harmful to the ligase activity. Beside, EDTA is not available in vivo.

f. It requires no input of energy. Energy is released by the joining of the sugar-phosphate backbone. That's incorrect because energy (ATP) is used to form the phosphodiester bond. Energy is required. Can't get around it..

h. A 3’ terminal phosphate is required. No. The 3' terminal of a deoxyrobose sugar is a hydroxyl group. The phosphate is found at the 5' end of the sugar.

i. T4 DNA ligase is most efficient at 20°C in vivo. No. It is most efficient at 25°C, not 20°C in vivo. But T4 ligase is effective at a wide range of temperature. For example, a typical temperature of 16-20°C is used overnight for blunt-end ligation.

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