Chapter 3 Nucleic Acid
1. Physical and chemical structure of DNA
Double-stranded helix
Major groove and minor groove
Base pairing
The two strands are antiparallel
G+C content (percent G+C)
Satellite DNA
Satellite DNA consists of highly repetitive DNA and is so called because repetitions of a short DNA sequence tend to produce a different frequency of the nucleotides adenine, cytosine, guanine and thymine, and thus have a different density from bulk DNA - such that they form a second or 'satellite' band when genomic DNA is separated on a density gradient.
2. Alternate DNA structure
Two bases have been extruded from base stacking at the junction. The white line goes from phosphate to phosphate along the chain. O is shown red, N blue, P yellow and C grey.
3. Circular and superhelical DNA
DNA can also form a double-stranded, covalently-closed circle. These circular molecules are often coiled into a superhelix, the formation of which is catalyzed by enzymes called topoisomerases.
4. Denaturation of DNA
Denaturation: A transition from the native to the denatured state
DNA denaturation: also called DNA melting, is the process by which double-stranded DNA unwinds and separates into single-stranded strands through the breaking of hydrogen bonding between the bases.
Hyperchromicity / Hyperchromic effect: the striking increase in absorbance of DNA (A260) caused by the denaturation of the double-stranded DNA molecule
Melting temperature (Tm) : the temperature at which half of the DNA strands are in the double-helical state and half are denatured. The melting temperature depends on both the length of the molecule, and the specific nucleotide position of that molecule.
Factors Affecting Tm
G-C content of sample
reagents that increase the solubility of the bases (anything that disrupts H-bonds or base stacking)
Salt concentration
pH
Length
5. Renaturation
Strands can be induced to renature (anneal) under proper conditions. Factors to consider:
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