2. Nucleic Acid Hybridization Theory

Question 1

Probe-‘patient’ NA target complexes that form

Can be specific or non-specific

Answer 1

Heteroduplex

Question 2

What happens when assay temp is ABOVE probe’s Tm

Answer 2

Denatured state favoured (no hybridization)

Question 3

Tm formula

Answer 3

Tm = 4(G+C) + 2(A+T)

Question 4

What is melting temperature (Tm)

Answer 4

Temperature at which HALF of a nucleic acid duplex is ds and HALF is ss (i.e. half of a duplex is melted)

Always lower than Td

Question 5

What is denaturation temperature (Td)

Answer 5

Temperature at which a double-stranded nucleic acid complex is COMPLETELY separated into single strands

Always higher than Tm

Question 6

What happens when assay temp is BELOW probe’s Tm

Answer 6

Specific AND non-specific hybridization occurs

Question 7

Formed when complementary probe strands or complementary genomic DNA strands re-nature

Answer 7

Homoduplex

Question 8

Ionic strength and temperature conditions (low/high) associated with HIGH stringency

Answer 8

LOW ionic strength

HIGH temperature