Genetics and Molecular Biology: DNA, RNA, Replication, and Transcription

Griffith experiment

Fred Griffith (1928): Worked with Streptococcus pneumoniae. Found two colony types: Smooth (S) - had a capsule and was more virulent. Rough (R) - lacked capsule and was nonvirulent. When heat-killed smooth cells were mixed with live rough cells, the live rough cells were transformed into smooth, virulent ones. Concluded there was a 'transforming factor' that changed rough into smooth.

Oswald Avery experiment

Oswald Avery et al. (1944): Selectively destroyed parts of S. pneumoniae cells (DNA, RNA, or protein) and repeated Griffith's experiment. Proteases → animals still died (protein not the factor). RNase → animals still died (RNA not the factor). DNase → animals lived (DNA destroyed). Conclusion: DNA held the key to transformation of rough to smooth — DNA is the genetic material.

Hershey-Chase experiment

DNA labeled with ³²P (Phosphorus) and Protein labeled with ³⁵S (Sulfur). Proved that: DNA, not protein, is the genetic material that enters the bacterial cell and carries genetic information.

Watson and Crick

Determine the structure of DNA (the double helix). Based on X-ray crystallography data from Wilkins and Rosalind Franklin (who did not receive credit).

Nitrogenous bases in RNA

Purines = Adenine (A) and Guanine (G); Pyrimidines = Cytosine (C) and Uracil (U); Base pairing: A-U and G-C.

Sugar in RNA

ribose

Comparison of DNA and RNA

Function: DNA - Genetic material; RNA - Carries the code for protein synthesis. Location: Bacteria - cytoplasm; Eukaryotes - nucleus; RNA - Cytoplasm. Bases: DNA - A, T, G, C; RNA - A, U, G, C. Sugar: DNA - Deoxyribose; RNA - Ribose. Structure: DNA - Double stranded; RNA - Single stranded.

Central dogma

Crick and Gomov (1957): Explained how DNA functions to make proteins. Central Dogma: DNA → RNA → Protein (Replication → Transcription → Translation).

Semi-conservative replication

One strand is used as a template to make a new strand. Each new DNA molecule has one old and one new strand.

Bidirectional replication

Two replication forks move in opposite directions around the circular bacterial chromosome.

Rolling circle replication

Seen in bacteriophages and during bacterial conjugation.

Steps of bacterial DNA replication

Helicase unwinds the DNA helix. Primase adds short RNA primers.

DNA Polymerase III

Adds nucleotides to synthesize the new strand.

DNA Polymerase I

Removes RNA primers and replaces them with DNA.

Ligase

Seals gaps in the backbone.

Topoisomerase

Relieves tension (supercoiling).

Direction of DNA Polymerase III

Reads the template strand 3′ → 5′ and synthesizes the new strand 5′ → 3′.

Leading Strand

Synthesized continuously toward replication fork.

Lagging Strand

Synthesized discontinuously away from fork.

Leading Strand Fragments

Single long strand.

Lagging Strand Fragments

Short Okazaki fragments.

Leading Strand Primers

One primer needed.

Lagging Strand Primers

Many primers needed.

Helicase

Unwinds the DNA helix.

Primase

Adds RNA primers.

DNA Polymerase III Role

Synthesizes new strand (5′→3′).

DNA Polymerase I Role

Removes primers and replaces with DNA.

Ligase Role

Seals nicks in the sugar-phosphate backbone.

Topoisomerase Role

Relieves supercoiling tension.

DNA Structure

Double stranded, contains thymine, sugar is deoxyribose, stores genetic info.

RNA Structure

Single stranded, contains uracil, sugar is ribose, carries out protein synthesis.

Transcription Initiation

RNA polymerase binds to the promoter region.

Transcription Elongation

RNA polymerase moves along DNA, adding nucleotides.

Transcription Termination

Stop signal ends transcription; mRNA is released.

mRNA

Carries codons to the ribosome.

rRNA

Forms part of ribosome structure and helps in translation.

tRNA

Carries anticodon and amino acid to the ribosome.

Codon

Groups of three nucleotides on mRNA that dictate which amino acid is added.

Anticodon

The complementary three-base sequence on tRNA.

Wobble

Only the first two nucleotides are required to encode the correct amino acid; the third base can vary without changing the amino acid.

Start Codon

AUG.

Stop Codons

UAA, UAG, UGA.

Promoter Location

DNA region where RNA polymerase binds to begin transcription.

Start Codon Location

On mRNA — signals translation to start.

A Site

Entry point for tRNA carrying amino acid.

P Site

Holds the growing polypeptide chain.

Bacteria Transcription & Translation

Occur simultaneously.

Eukaryotes Transcription & Translation

Occur separately.

mRNA Type in Bacteria

Polycistronic (multiple proteins from one mRNA).

mRNA Type in Eukaryotes

Monocistronic (one protein per mRNA).

Location of Transcription in Eukaryotes

Nucleus.

Location of Translation in Eukaryotes

Cytoplasm.

mRNA Processing in Bacteria

None.

mRNA Processing in Eukaryotes

Introns removed, exons spliced.