The two strand of DNA are anti-parallel. DNA is directional: one end, the 5' end, has a protruding phosphate group, while the other end, the 3' end, has a hydroxyl group., In prokaryotes, each cell has a single, large circular chromosome, as well as smaller circular pieces of DNA called plasmids., In eukaryotes, each cell has numerous large pieces of DNA called chromosomes. Organelles such as mitochondria and chloroplasts also contain DNA, Eukaryotic chromosomes can exist as chromosomes (condensed) or chromatin (dissfused)., Chromosome condensation involves tight wrapping of DNA around protein complexes called histones. Histones are positively charged and thus neutralise the negative charge on dna. The wrapping of a small stretch of dna around a histone core is called a nucleosome. When the nucleosome is secured with a histone clip (H1 histone), the structure is called a chromatosome. Tight packing of the nucleosomes/chromatosomes produces the 30 nm fibre. Further packing of the 30 nm fibre produces condensed chromosomes. Human chromosomes may be condensed 70,000 times., Techniques such as Spectral Karyotype analysis and Fluorescence In Situ Hybridisation (FISH) allow scientists to visualise chromosomes, chromosomal territories in nuclei and chromosomal changes., Chromosome binding dyes (Giemsa) allow chromosomal regions to be defines and chromosomal maps to be made.
Gene, Structure, Eukaryotic genes, Protein-coding region (open reading frame), Exon (protein coding segments), Intron (intervening regions - removed after transcription), Regulatory regions, 5' UTR (UnTranslated Region), Promoter, RNA polymerase binding site, Control regions (transcription fator binding sites), Proximal control sites (near the promoter), Distal control sites (far away from the promoter), Enhancer (stimulates transcription). Transcripton factors that bind to enhancers are called activators, Silencers (inhibit transcription). Transcription factors that bind to silencers are called repressors/suppressors, 3' UTR (UnTranslated Rgion), Termination Signals, Poly-adenylation signals (special sequence that results in a poly A tails being added to the mRNA after transcription), Prokaryotic genes, Operons, Regulatory regions, 5' UTR (UnTranslated Region), Promoter, RNA polymerase binding site, Operator, Repressor binding site, When repressor is bound to the operator, RNA polymerase is blocked from transcribing the structural genes (the operon is switched off). When operator is unoccupied, RNA polymerase is free to transcribe the structural genes (the operon is switched on)., Repressor protein, The repressor protein exists in two forms (allosteric). In one form, it is able to bind to the operator and the in the other form, it cannot bind to the operator. The shape is determined by the presence or absence of other molecules (metabolites)., Co-repressor, Tryptophan. The amino acid tryptophan binds to the trp repressor protein and changes its shape so that it binds to the operator (thus switching the trp operon off)., Lactose. The sugar lactose binds to the lac repressor protein and changes its shape so that it cannot bind to the operator (thus switching the lac operon off)., 3' UTR (UnTranslated Region), Termination Signals, Protein-coding region (open reading frame) - structural genes., All genes involved in a specific biochemical/molecular pathway are co-located., Expression, Central Dogma, The Central Dogma of molecular biology indicates that biological information that is sotred in nucleic acids, (DNA or RNA) is used to construct proteins. The flow of information is unidirections (nucleic acids to proteins). Information in RNA can be used to make DNA (this is called reverse transcription), but this is rare. Both DNA and RNA are capable of self-replication., Despite the many years since it was proposed, no exception to the Cental Dogma has been discovered., RNA viruses: capable of reverse transcription, but the synthesis of viral proteins involves unidirectional information flow from nucleic acids to proteins., Prions: prions do not replicate or carry out protein synthesis. Hence there is no 'need' for nucleic acids., RNA editing and RNA interference: these processes can either influence transcription or change the genetic information in RNA after transcription, but it does not change direction of the information flow from nucleic acids to proteins. There is no evidence that these processes change the genetic information in DNA., Prokaryotic genes, Operons, Inducible (Always OFF, unless turned on by inducer), Example: lac operon. When lactose is not present, repressor is bound to operator, preventing transcription by RNA polymerase (operon is OFF). In the presence of latose, allolactose (the inducer) leaks into the cell. It binds to the repressor protein, which changes its shape and falls off the DNA. RNA polymerase then transcribes the three genes in the lac operon, tunring turning the operon ON., Repressible (always ON unless turned off by repressor), Example, trp (tryptophan operon). Repressor protein cannot bind to the operator sequence in the DNA (operon if ON). However, when tryptophan is present in the cytoplasm, it binds to the repressor, causing it to change shape and bind to the operator. This prevents transcription by RNA polymerase, thus turning the operon OFF., mRNA is translated into a polypeptide chain by the ribosome complex. The ribosome complex consists of 2 subunit (small and large) and each subunit is held together by ribosomal RNA (rRNA). When assembled, the subunits create 3 pockets (E, P and A), Translation begins at the 5' end of the mRNA at a specific site known as the initiator codon (AUG, which specifies the amino acid, methionine). The ribosome moves along the mRNA, reading groups of 3 nucleotides (called codons). At each new codon, a tRNA bearing the appropriate amino acid enters the ribosome complex at the A site. A peptide bond is formed between the tRNAs at the P and A sites. This conitnues until the ribosome reaches the 3' end of the mRNA, which contains a stop codon. A molecule known as the release factor disassembles the ribosome, thus completing translation., Prokaryotic cells: consists of Archaea and Bacteria. Single celled organisms without organelles. Possess an external cell wall. Highly adaptible and lives in varied enviroments, including extremophiles, halophiles. Able to survive harsh conditions by sporulation., Ekaryotic genes, Transcription occurs in the nucleus. Transcription factors (activators) bind to distal contol elements (enhancers). This caused a bending protein to bind to and bend the DNA, so that the enhancer sequences come close the promoter region. General transcription factors bind to proximal control elements. This causes RNA polymerase to bind to the promoter. Transcription then proceeds as in the prokaryotes and is completed when the RNA polymerase recognises termination signals at the 3' UTR. Repressors are transcription factors that bind to suppressors (equivalent to enhancers, but with repressive function) to inhibit transcription., The mRNA that falls off the DNA is known as the pre-mRNA. This pre-mRNA has to be processed before it can act as a template for polypepide synthesis. The processing involves adding a cap structure at the 5' end, a poly-A tail at the 3' end, removal of introns (splicing) and occassionally, editing of the RNA sequences., mRNA is translated into a polypeptide chain by the ribosome complex. The ribosome complex consists of 2 subunit (small and large) and each subunit is held together by ribosomal RNA (rRNA). When assembled, the subunits create 3 pockets (E, P and A), Translation begins at the 5' end of the mRNA at a specific site known as the initiator codon (AUG, which specifies the amino acid, methionine). The ribosome moves along the mRNA, reading groups of 3 nucleotides (called codons). At each new codon, a tRNA bearing the appropriate amino acid enters the ribosome complex at the A site. A peptide bond is formed between the tRNAs at the P and A sites. This continues until the ribosome reaches the 3' end of the mRNA, which contains a stop codon. A molecule known as the release factor disassembles the ribosome, thus completing translation.If the polypeptide contains a signal sequence at its N-terminus, a signal recognition particle (SRP) binds to it and directs the entire ribosomal complex to the endoplasmic reticulum (ER). The newly synthesised polypeptides are released into the ER space (not always), where they are then modified by having sugars added to them (a process known as glycosylation; the polypeptides are known known as glycoproteins). The glycoproteins are then packaged into vesicles and sent to the Golgi Apparatus (GA), where further processing of the glycoproteins occur (e.g. further modifications of the sugars, addition of lipids, etc). Finally, the polypeptides are sorted and packaged into vesicles and sent to their final desitnations (e.g. plasma membrane)., Polypeptides have four levels of structure: Primary: linear sequence of amino acids; Secondary: hydrogen bonding within peprtide backbone to form alpha-helix or beta sheet; tertiary: three-dimentional structure of polypeptide that is stabilised by covalent (disulphide bonds), ionic and hydrophobic interactions; quaternary: association of multiple polypeptides. Polypeptides may consist of specialised regions called domains. Domains carry out speciliased functions, which are part of the overall function of the polypeptide., Protein Quantitation, The protein concentration in a sample can be determined using spectrophotometry. According to Beer's Law, the amount of light absorbed by any substance is directly proportional to its concentration in the solution. The absorbances of known concentrations of proteins are used to generate a standard curve and the absorbance of the 'unknown samples' is used to determine its protein content (using that standard curve)., Cells, Cells are the fundamental units of life. A unicellular organism, such as Paramecium, is a complete individual that is capable of performing all of life's functions., Feeding, Cytopharynx, food vacuole, cilia, Locomotion, Cilia, Osmoregulation, Contractile vacuole, Genetics, Macronucleus - diffused chromatin where gene expression occurs, Micronucleus - highly condensed chromosomes - used for reproduction, If proteins are misfolded, chaperone protein complexes will attempt to refold them., Misfolded proteins which cannot be repaired are destroyed in proteasomes., Differential gene expression, Since a transcription initiation complex must form before transcription can commence in eukaryotes, the presence/absence, as well as the types of transcription factors in a cell can determine which genes will be expressed., Different cell types contain different sets of transcription factors. Different genes will contain multiple control elements., This means that two different cell types in a single multicellular organism will express different sets of genes from the same genome. Hence, each cell type becomes functionally specialised because each will express a unique complement of proteins., The process of deriving unique cell types from undifferentiated embryonic cells is called differentation., Differentiation is the result of differential gene expression. During early embryonic development, cells migrate within the embryo. At this stage, the embryonic cells are undifferentiated and do not express 'cell type-specific' genes. When the cells arrive at their destinations, the switch on the expression of certain genes that will begin the process of differentiation. Initially, 'Master Regulatory Switches' are turned on. These are transcription factors. In turn, those transcription factors will induce the expression of 'differentiation' genes, thus driving the cells to their eventual developmental fates. MyoD is an example of a master regulatory switch, which in involved in the development of muscle cells.
Restriction enzymes are enzymes (mainly from prokaryotes) that cleave DNA in a sequence-specific manner. Each restriction enzyme recognises a specific DNA sequence, known as its restriction sequence or target sequence. The digested DNA has 'sticky' or 'blunt' ends., Enzymes called DNA ligases join DNA fragments together.
A method of separating DNA in an electric field. DNA is pushed through an agarose matrix by the electric field. DNA is separated according to size - small DNA fragments migrate further on a gel, compared to larger fragments. DNA is visualised by adding a fluorescent dye to the gel and then observing the gel under UV illumination.
A method of characterising differences in DNA using restriction enzymes. DNA from different sources are digested and then separated by electrophoresis. The similarity of the DNA profiles (number and sizes of fragments) indicates the degree of similarity or difference between the DNAs.
Cancer. A condition caused by the uncontrolled proliferation of cells. This occurs because of the breakdown of controls in the cell cycle.The direct causes of cancer are genetic mutations, although lifestyle and environmental factors alter the risk of developing cancer., Regulation: genetic causes., Signal to divide: Growth factors bind to cell surface receptors (usually tyrosine kinase receptors). This activates a signal transduction pathway that causes cells in G1 to enter the S phase. Mutations in certain genes, called proto-oncogenes, convert them into oncogenes. The proteins synthesised by oncogenes maintain the signal trasnduction pathway in an activated state, even though grwoth factors are not bound to the cell surface receptors. One example of an oncogene is Ras., The process of converting a normal cell into a cancer cell is called transformation. Multiple mutations, in both oncogenes and tumour suppressor genes, are required for this., Signal to stop dividing. Controls at the checkpoints ensure that the cells do not progress to the next stage of the cycle unless conditions and signals are appropriate. One group of such regulators are the tumour suppressors (e.g. p53). Mutations in the tumour suppressors can cause uncontrolled passage through the cell cycle.
Stages, M, Mitosis: DNA that was replicated in S phase is separated in mitosis. Mitosis occurs in somatic cells. Two daughter cells are produced from a parental cell and the daughter cells have identical genetic makeup to that of the parental cell., Prophase: Condensation of chromosomes. Spindle body replicates and moves to the opposite ends of the cell.Spindle body synthesises spindle fibres (microtubules). Spindle fibres from opposite sindle bodies attach to different sister chromatids. Nuclear membrane disintegrates., Metaphase. Chromosomes move towards the metaphase plate., Anaphase. Separation of sister chromatids to opposite ends of cell. Separation of chromatids occurs because of depolymerisation of spindle fibres., Telophase. Completion of chromosomal movements. Reformation of nuclear envelope. Disintegration of spindle fibres., Cytokinesis. Contraction of actin fibres just beneath the plasma membrane causes the membrane to move towads the cytoplasm. This mebrane contraction occurs between the separated nuclei. In plant cells, cytokinesis is followed by the laying down of a cellulose cell wall., Meiosis. Occurs in testes and ovaries durinng the formation of gametes. Results in the formation of 4 daughter cells from one parental cell. the genetic complement of the daughter cells is different to that of the parental cells. Genetic diversity occurs becuase of (i) crossing over of non-sister chromatids of homologous chromosomes - prophase I, (ii) random assortment of homologous chromosomes during metaphase I and (iii) random fertilisation of gametes during fertilisation. Sexual reproduction brings new combination of alleles together, which may bring about new properties in populations. It is thought that the evolution of immunological defense against rapidly evolving pathogen is one of the main advantages conferred by sexual reproduction., Prophase I: condensation of chromosomes. Crossing over of non-sister chromatids of homologous chromosomes. Spindle body duplicates and moves to opposite ends of the cell, while producing spindle fibres. pindle fibres from opposite ends conenct to different homologous chromosomes. Nuclear membrane disintegrates., Metaphase I. Alignment of chromosomes at the metaphase plate. Homologous chromosomes site side-by-side at the metaphase plate., Anaphase I: separation of homologous chromosomes (not chormatids)., Telophase I: reformation of nuclei. Nuclear membrane reforms, but spindle fibres disintegrates., Cytokinesis, Prophase II: same as mitosis prophase., Metaphase II: same as mitosis metaphase., Anaphase II: same as mitosis anaphase - separation of chromatids., Telophase II: same as mitosis telophase, Cytokinesis: formation of four daughter cells. In females, one of the two daughter cells at each cytokinesis degenerates into polar bodies. The cytoplasmic contents of these cells are transferred to the other daughter cell, leaving behind residual nuclei. These residual nuclei form highly refractile polar bodies. Only one daughter cell (egg) remains after one round of meiosis, G2, Cell recovers from S-phase. Cell contains twice the amount of DNA as it did in the G1 phase. At the G2/M checkpoint, the cell prepares for mitosis by synthesising proteins for that stage (spindle sibres, etc)., S, DNA replication: DNA is unwound by enzymes (DNA gyrase or DNA topoisomerase). DNA helicases open up the double helix at special sites called 'origins of replication' (which are kept apart by single stranded binding proteins (SSSB)). Primase synthesises a short RNA fragment (primer). DNA polymerase extends the primer towards the 3' end (this is the replication of the leading strand). For the other stand, primase synthesises a short primer slightly away from the origin of replication, which DNA polymerase extend towards the origin of replication. This strand is the lagging strand. The lagging strand is replicated as several short strand (called Okazaki fragments). Then, another form of DNA ploymerase replaces all RNA primers with DNA. the enzyme DNA ligase joins the backbones of the newly synthesised strands. Replicated DNAs remain attached together and each replicated molecule is referred to as a sister chromatid. This type of replication is known as semi-conservative replication., The 3' ends of the lagging strand are not completely replicated. This is because there is no site for the synthesis of RNA primers at the 3' ends. This causes the 3' ends to shorten everytime DNA replicates. Eventually, the 3' ends rode to the point where essential genes are deleted (critical length). This activates the p53 system, which stops the cell from undergoing further rounds of DNA replication and cell division (cell cycle). The cell exits the cell cycle and undergoes terminal differentiation and will die by apoptosis. A cell in this stage is said to be senescent. It means that cells have a finite lifetime (Hayflick limit, 40-70 divisions), before dying., In embryonic cells and stem cells, the ends of chromosomes are linked to a large protein complex known as telomerase. Telomerase contains an RNA fragment, from which DNA polymerase can extend and replicate the 3' ends. Hence, in these cell types, DNA is replicated without terminal erosion. At a certain strage of development, cells will not express telomerase. Terminal erosion occurs during DNA replication and the cell will eventually stop dividing., G1, Growth phase of the cell. At the G1/S checkpoint, the cell prepares for the S-phase by synthesisng the porteins required for DNA replication.
The sizes of genomes is unrelated to complexity.
Thus DNA carries the genetic information that specifies biological traits (e.g. pathogenic, non-pathogenic, etc).
This confirms that only DNA can be passed from parents (i.e. parental viruses in this example) to offspring (virions in this example). Thus it is the molecule of heredity.
Gene duplication provide extra copies of genes, which can acquire new mutations and diversify. The mutated forms of 'original' genes are referred to as alleles. Sometimes alleles confer new functions (e.g. a slightly different trait). New genes are also created by shuffling domain-encoding exons., Groups of sequences within a genome are referred to as markers and can be used to identify individuals. This allows us to trance the movement of DNA during evolution. Mitochondrial DNA enables us to trance our matrilineal ancestry, which Y-chromosome markers allows us to trance our partilineal ancestry (only in males). Such studies strongly support that modern humans arose in Africa and emigrated to other parts of the world. Prior to that, the lineage leading to the Neanderthals had left Africa for Europe and Asia. Although modern humans and the Neanderthals remained largely separate, molecular evidence suggests tjhat a small degree of intermingling did occur between these hominids. Molecular data has also provided information on the migration of large groups of peoples around the world in our history.
Two key aspects of scientific investigations are the development of hypotheses and theories. BOTH describe aspects of science - they are equally valide, but differ in scope., Scientific hypotheses are testable and falsifiable. Controlled experiments are used to the validity of hypotheses, Scientific approaches often use either inductive or deductive reasoning.
The basic ideas of evolution are (1) biological variation of traits exist in populations and are inherited; (2) changes in the environment or competition may favour individuals with specific traits over others; (3) individual possessing favourable traits are better adapted to survive and produce more offspring than others, thus producing a change in the structure of populations., There is overwhelming evidence for evolution as a biological process: fossils, biogeography, molecular evidence, etc.
Types of mutations, Chromosomal mutations. Insertions, deletions, inversion, deletions, translocations, Point mutations. Silent, Missense, Nonsense, Frameshift (substitutions,insertions and deletions)
Effects of mutations, Gain-of-function mutations: confers new biological functions, Loss-of-function mutations: Loss of biological function.
Repair, Cells contain numerous mechanisms to repair damaged DNA, including excision repair
Cleavage: rapid mitotic division of zygote to produce an embryo with a large numer of cells. Cells in the interior die by apoptosis, resulting in a fluid filled interior. this is the blastula stage., Cellular migration. Also known as gastrulation. Results in the formation of tissue layers (endoderm, mesoderm and ectoderm)., Laying down of the body axes (anterio-posterior/dorso-ventral). Body plan/patterning/positioning. Cellular migration into the interior of the embryo is guided by cytoplasmic protein gradients, called morphogen gradients. A cell knows where it is located within the embryo by determining the concentration of different morphogens it is exposed to. Depending on its locations within the embryo, cells will turn on the expression of homeobox genes (Hox genes) that are specific to the embryonic regions they are located in., Differentiation: Depending on the set of Hox gene that they express, cell will turn off the expression of differentiation genes. E.g., myoblasts (muscle stem cells) will turn on the expression of the MyoD gene, which in turn induces the expression of actin and myosin genes. This converts the myoblast into a differentiated muscle cell., Apoptosis. Differentiation also intitates processes that will lead to the orderly death of the cell (apoptosis). Cells will initally express survival genes (bcl-2), which inhibit the apoptotic pathway. When the expression of survival genes is turned off, apotosis will kill the cell. Apoptosis is an essential part of multicellular development and is important for processes such as the development of body shape.