分子生物学(英文版)

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 sequence composition 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:

● Temperature

● Salt concentration

● DNA concentration

● Time

Repetitive Sequences

●Unique: Single Copy Genes

● Slightly repetitive (2-10 copies)

● Middle repetitive (10- hundreds)

--Clustered

--Dispersed

Highly repetitive (hundreds to millions)

--Short sequences in satellite DNA

--Sequences of normal length in certain genes that exist in very large numbers

C-value Paradox

There is apparently a lack of association between C-value (the amount of DNA present in the haploid genome of different organisms )and the degree of organismal complexity of various multi-cellular organisms. In 1971, Thomas named this phenomenon, “C-value Paradox”.

在每一种生物中其单倍体基因组的DNA总量是特异的，被称为C值 (C Value)。 C值和生物结构或组成的复杂性不一致的现象称为C值悖论(C-value paradox)。

6. Hybridization

Hybridization: the technique wherein renatured DNA is formed from separate single-stranded samples . Heteroduplexing: renaturation combined with electron microscopy in a procedure allows the localization of common, distinct,and missing sequences in DNA.

DNA-RNA hybridization (Northern hybridization): the use of filter hybridization to detect sequence complementarity between a single strand of DNA and an RNA molecule.

7. The structure of RNA

Types: mRNA, tRNA, rRNA

Distinctions:

- ribose replaces deoxyribose;

- U replaces T;