9-1 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Chapter 9: Genes, chromosomes and DNA

9-2 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Tracking the genetic material 1869—chromatin isolated by Miescher, containing nucleic acid and protein Chromosomes consist of DNA and proteins 1900—concept of ‘Mendelian inheritance’ controlled by ‘genes’ 1910—Morgan and others noted parallel inheritance of ‘genes’ with chromosomes, suggesting that genes were ‘on’ the chromosomes (cont.)

9-3 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Tracking the genetic material (cont.) The transforming principle in Streptococcus pneumoniae, where virulence can be transferred by cellular extracts containing DNA (Avery, McLeod & McCarty 1944) –mice injected with live non-virulent bacteria and heat- killed virulent bacterial material died –neither preparation on its own killed the mice –non-virulent strain was ‘transformed’ by the virulent material –the virulence acquired from the heat-killed strain was passed on to progeny of the transformed bacteria (cont.)

9-4 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Fig. 9.5: Transforming principle in Streptococcus pneumoniae

9-5 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Tracking the genetic material (cont.) DNA, not protein, is the genetic information (Hershey & Chase 1952) –bacteriophage DNA or protein was specifically radioactively labelled –bacteriophage infected bacteria—new bacteriophage produced by infected organisms –the presence of radiolabel inside infected bacteria was only detected when the DNA was radiolabelled –no radiolabelled protein was found inside the bacteria

9-6 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Fig. 9.6: Radioactive labelling of DNA with 32 P or protein with 35 S

9-7 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Chromosomes DNA is organised into chromosomes Each chromosome is a single DNA molecule In eukaryotic cells, chromosomes are located in the nucleus Each species has a unique chromosome complement—shape, size and number Centromere essential for segregation during cell division

9-8 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Fig. 9.1: Stained human chromosomes

9-9 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Chromosome structure Multiple levels of DNA folding –nucleosome: 146 base pairs (bp) are coiled in 1.75 turns around a core of histone proteins (H2A, H2B, H3, H4) 10 nm diameter (cont.)

9-10 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Fig. 9.3: Model of a nucleosome particle

9-11 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Chromosome structure (cont.) This string of nucleosome ‘beads’ is then further coiled into chromatin fibres 30 nm diameter Metaphase chromosomes are further condensed to about 1/ of their full length Loops of 20–100 kb are attached to a central protein scaffold

9-12 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Fig. 9.4: A condensed chromosome in metaphase

9-13 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint DNA structure DNA is a double-stranded molecule twisted into a helix Each strand, comprising a sugar-phosphate backbone and attached bases, is connected to a complementary strand by non-covalent hydrogen bonding between paired bases The bases are adenine (A), thymine (T), cytosine (C) and guanine (G) (cont.)

9-14 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint DNA structure (cont.) DNA consists of four different nucleotides Each nucleotide has three parts: a phosphate group, a pentose sugar and an organic base (cont.)

9-15 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Fig. 9.7: Molecular structure of DNA

9-16 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint DNA structure (cont.) Bases are purines (A and G) and pyrimidines (C and T) Purines have a pair of fused rings; pyrimidines only have one A and T are connected by two hydrogen bonds; G and C are connected by three hydrogen bonds The number of bonds is the basis of specific pairing between the bases (cont.)

9-17 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint DNA structure (cont.) Nucleotides are linked together by phosphodiester bonds Nucleic acids have distinct ends –the 3’ end has a free hydroxyl group on the 3’ carbon of a sugar –the 5’ end has a free phosphate group at the 5’ carbon of the sugar The two strands of the helix are antiparallel: the 5’ end of one strand is directly apposed to the 3’ end of the other strand

9-18 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint DNA replication DNA is replicated semi-conservatively—each separate strand provides the template for new strand synthesis by the base-pairing rules Semi-conservative replication allows synthesis of new strands with high fidelity New DNA molecules consist of one ‘old’ strand from the original molecule and one newly synthesised strand

9-19 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Fig. 9.8a: Semiconservative replication

9-20 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Fig. 9.8b: Sequence-based representation of replicating DNA

9-21 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint DNA replication in prokaryotes Bacteria have a single circular chromosome Replication begins at a single origin of replication A nick is made in at least one strand and the molecule unwinds A replication fork is formed on each side of the origin as small lengths of DNA separate for synthesis of new strands The two replication forks eventually meet at the terminus

9-22 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Fig. 9.10: DNA synthesis in circular chromosomes

9-23 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Enzymes in replication Requires gyrases to unwind the supercoiled helices and helicases to separate the strands New strand synthesis is performed by DNA polymerases –DNA polymerase III attaches bases in the 5’  3’ direction –DNA polymerase I checks the added base and corrects it by 3’ to 5’ exonuclease activity—also removes RNA primers used to initiate replication DNA polymerases require priming to initiate strand extension –a short RNA primer with a 3’ OH group is added to the template strand by a primase (cont.)

9-24 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Fig. 9.13: Initiation of DNA synthesis

9-25 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Enzymes in replication (cont.) Synthesis always proceeds 5’  3’ on the strand being produced therefore –one strand is synthesised continuously (leading strand) –the other (lagging strand) is synthesised discontinuously as the replication fork moves along the template strand –primases attach a series of primers along the template strand –DNA polymerase extends the primers away from the replication fork –the resulting Okazaki fragments are then ligated by DNA ligase

9-26 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Fig. 9.11: Replication fork of Escherichia coli

9-27 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Replication in eukaryotes Chromosomes have many origins of replication Two replication forks are formed at each origin Synthesis proceeds 5’ to 3’ at each unit of replication (replicon) with leading and lagging strands (cont.)

9-28 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Fig. 9.14: DNA synthesis in a chromosome of a eukaryote

9-29 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Replication in eukaryotes (cont.) Okazaki fragments are shorter than in prokaryotes Leading and lagging strand synthesis in human cells is performed by different DNA polymerases Multiple replicons are necessary due to the large size of eukaryote chromosomes Replicons are initiated at different times –chromosomes have early-, mid- or late-replicating regions –gene-rich regions tend to be replicated first

9-30 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Telomeres during replication DNA polymerases only replicate DNA 5’ to 3’ and need a primer When the primer is removed from the 5’ end of the new strand a gap is left from which DNA polymerase cannot extend At each round of cell division chromosomes would become shorter (cont.)

9-31 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Telomeres during replication (cont.) To overcome this problem –chromosomes have telomeres repeat DNA sequences up to 10–15 kb –added to chromosome ends by telomerase –priming provided by RNA molecule within the telomerase complex –chromosome length is maintained (cont.)

9-32 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Fig. 9.15: Completion of replication at ends (telomeres) of eukaryotic chromosomes

9-33 Copyright  2005 McGraw-Hill Australia Pty Ltd PPTs t/a Biology: An Australian focus 3e by Knox, Ladiges, Evans and Saint Telomeres during replication (cont.) Mammalian somatic cells have no telomerase activity so become shorter with age This limits the number of divisions each cell can undergo Essential sequences are eventually lost and the cell dies Restoration of telomerase activity allows cells to proliferate indefinitely Telomerase is important in ageing and cancer