The \"melting\" or dissociation temperature for short DNA probes of lessthanoreq

Question

The "melting" or dissociation temperature for short DNA probes of lessthanorequalto 20 nucleptides is sometimes estimated using the equation below. T_m (degree C) = (4degree)(no. of GC pair) + (2degree)(no. of AT pair) - (5degree)(no. of mismatches) A. Estimate the T_m of the probe from its exactly complementary sequence: I) ATGCATGCATGCATGCAT (Ignore the underlining until part B.) II) GCATGCATGCA TGCATGC III) GCGCGCGCGC B. Probe (I) above is used to detect a mutated sequence in which the region complementary to the underlined 5 'ATG3' is not the expected 3 'TAC5' but rather 3 'CAT5'. (All other positions in the mutated sequence match the probe exactly). Estimate T_m for the probe (I) binding to this mutated sequence.

Explanation / Answer

A. As per the formula,

Tm = 4 (G+C) + 2 (A+T) Since the pairing is between exactly complementary sequence, there will be 0 mismatches

Hence,

i) Tm = 4 x 8 + 2 x 10= 32+20 = 52 degrees. ( there are 8 G+C and 10 A+T)

ii) Tm = 4 x 10 + 2 x 8 = 40 + 16 = 56 degrees (there are 10 G+C and 8 A +T)

iii) Tm = 4 x 10 + 2 x 0 = 40 degrees

B. As per the description given in the question, Tm for the mutated probe will be calculated as follows:

Tm = 4 x (G + C) + 2 (A +T)- 5 (No. mismatches

Now if the complementary sequence for ATG is CAT (instead of the perfect match TAC), tger are two mismatches (A-C nd G-T). Hence,

Tm = 4 x 7 + 2 x 9 - 5 x 2= 28 + 18 -10

= 28 + 18 -10

= 46-10

= 36

Related Questions

Navigate

• Browse (All)
• Browse T
• Subjects

---

---

Integrity-first tutoring: explanations and feedback only — we do not complete graded work. Learn more.