History and Structure of DNA

History of the Gene

• DNA was first described in 1869 by Johannes Miescher
• Frederick Griffith in 1928, observed harmless bacteria transformed into lethal ones
• Hereditary information must be transmitted in the molecule
• Hereditary material encodes information required to build a new individual
• Proteins and DNA were candidates for hereditary material
• Avery MacLeod and McCarty in 1944, confirmed that the “transforming principle" was a nucleic acid
• In 1952, Hershey and Chase established that DNA transmits hereditary information

Discovery of the Structure of DNA

• The DNA molecule is a polymer of nucleotides
• Each nucleotide is composed of a nitrogenous base, a five-carbon sugar (deoxyribose), and a phosphate group
• There are four nitrogenous bases in DNA: adenine, guanine, cytosine, and thymine
• Each DNA nucleotide consists of a sugar, a phosphate group, and a base
• Cytosine and thymine are pyrimidines, while guanine and adenine are purines
• Erwin Chargaff's rules (1950) include: The amount of Adenine equals Thymine, and Guanine equals Cytosine
• Each species differs in its Adenine to Guanine ratios
• In the 1950s, Rosalind Franklin made an x-ray diffraction image of DNA
• James Watson and Francis Crick proposed the double-helix structure model of DNA
• DNA molecule is composed of two strands
• Each strand contains nucleotides covalently bonded together between the phosphate group and the deoxyribose sugar of the next
• The bases of one strand bond to the bases of the second strand with hydrogen bonds
• Adenine always bonds with thymine, and cytosine always bonds with guanine
• This bonding causes the two strands to spiral around each other: double helix
• Ribonucleic acid (RNA) is a second nucleic acid found in cells
• Unlike DNA, RNA is a single-stranded polymer of nucleotides
• RNA contains the sugar ribose, not deoxyribose, and the nucleotide uracil, not thymine
• RNA molecules function in the process of forming proteins from the genetic code in DNA
• Prokaryotes contain a single, double-stranded circular chromosome
• Eukaryotes contain double-stranded linear DNA molecules packaged into chromosomes
• The DNA helix is wrapped around histone proteins to form nucleosomes
• During mitosis and meiosis, the protein coils become further coiled to facilitate chromosome movement

DNA Replication

• DNA replicates by a semi-conservative method, acting as a template for new DNA to be synthesized
• After replication, each DNA has one parental (old) and one daughter (new) strand
• Replication in eukaryotes starts at multiple origins of replication
• The DNA is opened with enzymes that form the replication fork
• Primase synthesizes an RNA primer to initiate synthesis by DNA polymerase, which can only add nucleotides in one direction
• One strand is synthesized continuously in the direction of the replication fork, also called the leading strand
• The other strand, also called the lagging strand, is synthesized away from the replication fork in short stretches of DNA
• Once replication is completed, the RNA primers are replaced by DNA nucleotides and the DNA is sealed with DNA ligase
• The ends of eukaryotic chromosomes pose a problem since polymerase cannot extend them without a primer
• Telomerase, an enzyme with an inbuilt RNA template, extends the ends by copying and extending one end of the chromosome
• DNA polymerase extends the DNA using the primer, protecting the telomeres of the chromosomes
• Elizabeth Blackburn discovered how telomerase works

Mutations

• Cells have mechanisms for repairing DNA damage or errors in replication
• Mismatch repair replaces nucleotides paired with non-complementary bases
• Nucleotide excision repair removes damaged bases, such as thymine dimers

Central Dogma of Molecular Biology

• The central dogma describes the flow of genetic information from genes to mRNA to proteins
• Genes are used to make mRNA during transcription
• mRNA is used to synthesize proteins during translation

DNA Transcription

• Initiation of mRNA synthesis relies on when RNA polymerase binds to the promoter sequence on the DNA template
• Elongation involves RNA polymerase synthesizing new mRNA
• Termination frees the mRNA and occurs by mechanisms that stall the RNA polymerase and cause it to fall off the DNA template
• Newly transcribed eukaryotic RNAs are modified with a cap and a poly-A tail
• These structures protect the mature mRNA from degradation and help export it from the nucleus
• Eukaryotic mRNAs also undergo splicing: introns are removed and exons are reconnected with single-nucleotide accuracy
• Only the finished mRNAs are exported from the nucleus to the cytoplasm

Translation

• The genetic code is the correspondence between the three-nucleotide mRNA codon and an amino acid
• The genetic code is translated by the tRNA molecules, which associate a specific codon with a specific amino acid
• There are 64 triplet codons in mRNA which specify only 20 amino acids and three stop codons, this makes the genetic code degenerate
• Every species on the planet uses the same genetic code
• Translation players include the mRNA template, ribosomes, tRNAs, and enzymatic factors
• A ribosome binds to the mRNA template
• Translation begins at the initiating AUG on the mRNA
• The formation of bonds occurs between sequential amino acids specified by the mRNA template
• Ribosomes accept charged tRNAs and catalyze the bonding between the new amino acid and the growing polypeptide
• When a stop codon is encountered, the components dissociate and free the new protein

Gene Regulation

• Somatic cells within an organism contain the same DNA, but do not express the same proteins
• Proteins are expressed only when they are needed
• In each cell type, the protein type and amount are regulated by controlling gene expression
• Prokaryotic cells use operons which contain multiple genes under the control of one promoter
• Eukaryotic cell gene expression is regulated at the epigenetic, transcriptional, post-transcriptional, translational, and posttranslational levels
• Eukaryotic gene expression is regulated during transcription and RNA processing, which occur in the nucleus
• Protein translation, which occurs in the cytoplasm, is also involved in gene regulation
• Post-translational modifications of proteins may further influence this process

Biotechnology

• Molecular and cellular techniques of biotechnology allow researchers to genetically engineer organisms and modify them to achieve traits
• Nucleic acids can be isolated from cells for analysis by breaking open the cells and destroying other major macromolecules
• DNA can be cut subsequently and re-spliced using restriction enzymes
• Fragmented or whole chromosomes can be separated based on size by gel electrophoresis
• Short stretches of DNA can be amplified by PCR

Cloning

• Dolly the sheep was the first cloned mammal
• Dolly was created when a nucleus was removed from a donor egg cell and the enucleated egg was placed and shocked in order to fuse with another cell
• The cells were shocked again to start division and were allowed to divide for several days until an early embryonic stage was reached, before being implanted in a surrogate mother
• Cloning may involve cloning small DNA fragments (molecular cloning) or cloning entire organisms (reproductive cloning)
• In reproductive cloning, a donor nucleus is put into an enucleated egg cell, which is then stimulated to divide and develop into an organism
• In molecular cloning with bacteria, a desired DNA fragment is inserted into a bacterial plasmid using restriction enzymes, the plasmid is taken up by a bacterium, and then it will express the foreign DNA

Genetically Modified

• Through other techniques, foreign genes may be inserted into eukaryotic organisms: called transgenic organisms
• In reverse genetics methods, a gene is mutated or removed to identify its effect on the phenotype of the whole organism and determine its function

Genomics

• Genome mapping provides an outline for the location of genes within a genome
• Whole genome sequencing is the latest available resource to treat genetic diseases
• The most detailed information is available through sequence mapping
• Information from all mapping and sequencing sources is combined to study an entire genome