9-1Copyright 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

9-2Copyright 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.)

9-3Copyright 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.)

9-4Copyright 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

9-5Copyright 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

9-6Copyright 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 32P or protein with 35S

9-7Copyright 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

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Fig. 9.1: Stained human chromosomes

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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.)

9-10Copyright 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

9-11Copyright 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 1/10 000 of their full length

• Loops of 20–100 kb are attached to a central protein scaffold

9-12Copyright 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

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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.)

9-14Copyright 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.)

9-15Copyright 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

9-16Copyright 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.)

9-17Copyright 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

9-18Copyright 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

9-19Copyright 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

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Fig. 9.8b: Sequence-based representation of replicating DNA

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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

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Fig. 9.10: DNA synthesis in circular chromosomes

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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.)

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Fig. 9.13: Initiation of DNA synthesis

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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

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Fig. 9.11: Replication fork of Escherichia coli

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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.)

9-28Copyright 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

9-29Copyright 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

9-30Copyright 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.)

9-31Copyright 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.)

9-32Copyright 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

9-33Copyright 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