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1:​ ​DNA,​ ​CHROMOSOMES,​ ​AND​ ​GENETICS

2

2:​ ​TRANSCRIPTION

4

2:​ ​POST-TRANSCRIPTIONAL​ ​MODIFICATION

4

2:​ ​RNA​ ​VS
...
​ ​all​ ​of​ ​the​ ​DNA​ ​found​ ​in​ ​a
body​ ​cell)
● Genomics:​ ​the​ ​genome-wide​ ​analysis​ ​of​ ​gene​ ​structure​ ​and​ ​expression
○ Genomes​ ​can​ ​be​ ​entirely​ ​sequenced​ ​(eg
...
​ ​the​ ​human​ ​genome​ ​is:
○ 21%​ ​LINEs
○ 13%​ ​SINEs
○ 8%​ ​retroviral-like​ ​elements
○ 3%​ ​DNA-only​ ​transposon​ ​“fossils”
○ 3%​ ​segmental​ ​duplications
○ 5%​ ​simple​ ​sequence​ ​repeats
○ 37
...
5%​ ​exons​ ​(protein-coding​ ​regions)
○ 8%​ ​heterochromatin
● The​ ​size​ ​of​ ​a​ ​genome​ ​is​ ​not​ ​directly​ ​related​ ​to​ ​its​ ​biological​ ​complexity
○ The​ ​bulk​ ​of​ ​genomes​ ​are​ ​heterochromatin​ ​(not​ ​sequenced)​ ​and​ ​introns/other
non-coding​ ​parts​ ​of​ ​genes​ ​from​ ​euchromatin
○

Joanna​ ​Griffith​ ​(2017)

Comparisons​ ​of​ ​genomes​ ​among​ ​species​ ​can​ ​suggest​ ​evolutionary​ ​and​ ​functional
relationships​ ​among​ ​genes
--------------------------------------------------------------------------------------------------------------------------●

2:​ ​TRANSCRIPTION
Genetic​ ​information​ ​must​ ​be​ ​stable​ ​for​ ​storage,​ ​but​ ​also​ ​available​ ​to​ ​direct​ ​cellular
processes
○ The​ ​genetic​ ​instructions​ ​carried​ ​by​ ​the​ ​DNA​ ​must​ ​be​ ​transcribed​ ​into​ ​RNA
■ Messenger​ ​RNA​ ​(mRNA)​ ​acts​ ​as​ ​a​ ​“messenger”​ ​to​ ​direct​ ​the
production​ ​of​ ​proteins
● Transcription​ ​produces​ ​an​ ​RNA​ ​molecule​ ​that​ ​is​ ​complementary​ ​to​ ​one​ ​strand​ ​of
DNA
● RNA​ ​is​ ​synthesised​ ​in​ ​a​ ​5’​ ​to​ ​3’​ ​direction​ ​from​ ​a​ ​DNA​ ​template​ ​by​ ​RNA​ ​polymerase
● How​ ​does​ ​transcriptional​ ​machinery​ ​know​ ​where​ ​to​ ​start​ ​and​ ​stop?
○ ATG​ ​start​ ​codon
○ Stop​ ​codons​ ​can​ ​differ
● Initiation​ ​of​ ​transcription​ ​(general​ ​principles):
○ RNA​ ​polymerase​ ​interacts​ ​with​ ​transcription​ ​factors​ ​when​ ​it​ ​binds​ ​to​ ​the
promoter​ ​region​ ​of​ ​the​ ​DNA
○ A​ ​basic​ ​promoter​ ​is​ ​required​ ​for​ ​RNA​ ​polymerase​ ​to​ ​bind​ ​and​ ​initiate
transcription​ ​at​ ​the​ ​appropriate​ ​site
■ Additional​ ​control​ ​sequences​ ​can​ ​determine​ ​when​ ​a​ ​gene​ ​is
transcribed
What​ ​types​ ​of​ ​RNA​ ​are​ ​transcribed​ ​from​ ​DNA?
● Ribosomal​ ​RNA​ ​(rRNA)
○ Forms​ ​part​ ​of​ ​the​ ​ribosome,​ ​catalyses​ ​protein​ ​synthesis
● Messenger​ ​RNA​ ​(mRNA)
○ Encodes​ ​proteins
● Transfer​ ​RNA​ ​(tRNA)
○ Acts​ ​in​ ​protein​ ​synthesis​ ​as​ ​adaptors​ ​between​ ​specific​ ​codon​ ​sequences​ ​on
the​ ​mRNA​ ​and​ ​amino​ ​acids
● Small​ ​RNA​ ​(small​ ​nuclear​ ​RNA,​ ​snRNA)
○ Used​ ​in​ ​pre-mRNA​ ​processing,​ ​transport​ ​of​ ​proteins​ ​to​ ​the​ ​endoplasmic
reticulum,​ ​and​ ​other​ ​cellular​ ​processes
● Micro​ ​RNA​ ​(miRNA)
○ Act​ ​in​ ​regulation​ ​of​ ​gene​ ​expression,​ ​eg
...
​ ​DNA
RNA​ ​has​ ​a​ ​ribose​ ​sugar,​ ​DNA​ ​has​ ​deoxyribose
RNA​ ​has​ ​uracil,​ ​DNA​ ​has​ ​thymine
RNA​ ​is​ ​chemically​ ​more​ ​reactive​ ​than​ ​DNA
○ Ribose​ ​has​ ​two​ ​OH​ ​groups
● RNA​ ​is​ ​less​ ​stable​ ​than​ ​DNA
○ Does​ ​not​ ​last​ ​as​ ​long
■ Can​ ​only​ ​be​ ​used​ ​to​ ​synthesise​ ​proteins​ ​for​ ​a​ ​short​ ​amount​ ​of​ ​time
○ DNA​ ​can​ ​be​ ​stored​ ​and​ ​used​ ​for​ ​many​ ​years
● RNA​ ​is​ ​more​ ​prone​ ​to​ ​mutate​ ​than​ ​DNA
○ Cytosine​ ​deamination​ ​to​ ​uracil​ ​cannot​ ​be​ ​detected​ ​and​ ​repaired​ ​in​ ​RNA
● RNA​ ​is​ ​single-stranded,​ ​DNA​ ​is​ ​double-stranded
● RNA​ ​can​ ​base​ ​pair​ ​to​ ​form​ ​3D​ ​shapes​ ​(such​ ​as​ ​enzymes​ ​with​ ​enzymatic​ ​functions),
DNA​ ​cannot
--------------------------------------------------------------------------------------------------------------------------●
●
●

3:​ ​CRACKING​ ​THE​ ​GENETIC​ ​CODE
The​ ​genetic​ ​code​ ​is​ ​degenerate​ ​and​ ​a​ ​triplet​ ​code
● Degenerate:​ ​a​ ​single​ ​amino​ ​acid​ ​can​ ​be​ ​coded​ ​for​ ​by​ ​a​ ​number​ ​of​ ​different​ ​variations
of​ ​bases
● How​ ​many​ ​bases​ ​correspond​ ​to​ ​one​ ​amino​ ​acid?
○ Four​ ​different​ ​types​ ​of​ ​base​ ​in​ ​nucleic​ ​acids​ ​(A,​ ​C,​ ​T,​ ​G)
○ Twenty​ ​different​ ​types​ ​of​ ​amino​ ​acid​ ​in​ ​proteins
○ 1​ ​base​ ​per​ ​amino​ ​acid​ ​=​ ​only​ ​4​ ​possible​ ​amino​ ​acids
○ 2​ ​bases​ ​per​ ​amino​ ​acid​ ​=​ ​only​ ​16​ ​possible​ ​amino​ ​acids
○ 3​ ​bases​ ​per​ ​amino​ ​acid​ ​=​ ​64​ ​possible​ ​amino​ ​acids
■ Therefore,​ ​there​ ​must​ ​be​ ​3​ ​bases​ ​per​ ​amino​ ​acid​ ​(triplet​ ​code)​ ​and
more​ ​than​ ​one​ ​combination​ ​of​ ​three​ ​can​ ​code​ ​for​ ​one​ ​amino​ ​acid
(degenerate​ ​code)
● Crick,​ ​Brenner​ ​et​ ​al
○ Demonstrated​ ​the​ ​triplet​ ​code
○ Used​ ​the​ ​T4​ ​bacteriophage​ ​which​ ​infects​ ​E
...

coli​ ​that​ ​could​ ​be​ ​identified)
● Proflavin​ ​is​ ​a​ ​planar​ ​molecule​ ​that​ ​inserts​ ​between​ ​base​ ​pairs
in​ ​DNA​ ​and​ ​causes​ ​“frameshift”​ ​mutations​ ​(eg
...

● Non-overlapping:​ ​ABC,​ ​DEF,​ ​GHI,​ ​etc
...
​ ​control​ ​of​ ​iron​ ​import​ ​into​ ​human​ ​cells
○ Iron​ ​(Fe)​ ​binds​ ​to​ ​an​ ​extracellular​ ​protein​ ​called​ ​transferrin,​ ​and​ ​the
Fe-transferrin​ ​complex​ ​then​ ​enters​ ​the​ ​cell​ ​via​ ​the​ ​transferrin​ ​receptor
■ An​ ​‘Iron​ ​Response​ ​Element’​ ​on​ ​mRNA​ ​is​ ​recognised​ ​and​ ​bound​ ​by​ ​an
IRE​ ​binding​ ​protein
Methylation
● Methylating​ ​the​ ​5’​ ​end​ ​of​ ​a​ ​gene​ ​changes​ ​the​ ​levels​ ​of​ ​expression​ ​(on/off)
○ Methylating​ ​a​ ​mammal​ ​gene​ ​turns​ ​off​ ​the​ ​gene
○ Methylating​ ​an​ ​insect​ ​gene​ ​turns​ ​on​ ​the​ ​gene
--------------------------------------------------------------------------------------------------------------------------■

5:​ ​DNA​ ​REPLICATION
Semiconservative​ ​(Meselson​ ​and​ ​Stahl,​ ​1958)
DNA​ ​is​ ​replicated​ ​through​ ​complementary​ ​base​ ​pairing
Replication​ ​takes​ ​place​ ​5’​ ​to​ ​3’
DNA​ ​polymerase​ ​must​ ​have​ ​3’-OH​ ​residue​ ​to​ ​extend​ ​from
The​ ​breakage​ ​of​ ​phosphoanhydride​ ​bonds​ ​of​ ​dNTPs​ ​provides​ ​energy​ ​for
polymerisation
DNA​ ​replication​ ​is​ ​bidirectional​ ​from​ ​origins​ ​of​ ​replication
● Forms​ ​replication​ ​bubbles
● Replication​ ​fork
○ both​ ​strands​ ​(lagging​ ​and​ ​leading)​ ​are​ ​copied​ ​at​ ​replication​ ​forks,​ ​in​ ​a​ ​5’​ ​to​ ​3’
direction
■ Synthesis​ ​of​ ​the​ ​leading​ ​strand​ ​is​ ​continuous
■ Synthesis​ ​of​ ​the​ ​lagging​ ​strand​ ​is​ ​discontinuous,​ ​leaving​ ​Okazaki
fragments​ ​that​ ​are​ ​later​ ​joined​ ​by​ ​DNA​ ​ligases
● Enzymes​ ​at​ ​the​ ​replication​ ​fork
○ Helicase,​ ​unwinds​ ​the​ ​double​ ​helix​ ​of​ ​DNA
○ Ssbinding​ ​protein,​ ​stabilises​ ​ssDNA
○ Primase​ ​(RNA​ ​polymerase),​ ​synthesises​ ​RNA​ ​primers
○ Initiating​ ​DNA​ ​polymerase,​ ​synthesis​ ​the​ ​new​ ​DNA​ ​strand
○ Progressive​ ​DNA​ ​polymerase,​ ​proofreads​ ​the​ ​replicated​ ​strand
●
●
●
●
●

Joanna​ ​Griffith​ ​(2017)

○ Sliding​ ​clamp,​ ​keeps​ ​DNA​ ​polymerase​ ​on​ ​the​ ​DNA
○ Clamp​ ​loader,​ ​loads​ ​the​ ​sliding​ ​clamp​ ​and​ ​DNA​ ​polymerase​ ​onto​ ​the​ ​DNA
○ Nucleases,​ ​trim​ ​the​ ​Okazaki​ ​fragments
○ DNA​ ​ligases,​ ​join​ ​the​ ​Okazaki​ ​fragments
○ Replisome,​ ​other​ ​factors
● Speed​ ​of​ ​DNA​ ​replication
○ About​ ​50​ ​base​ ​pairs/second​ ​at​ ​every​ ​fork​ ​in​ ​eukaryotes
○ About​ ​1000​ ​base​ ​pairs/second​ ​at​ ​each​ ​fork​ ​in​ ​prokaryotes
■ High​ ​processivity​ ​because​ ​DNA​ ​polymerase​ ​attaches​ ​to​ ​a​ ​sliding
clamp
DNA​ ​replication​ ​through​ ​PCR
● Needed:
○ Enzyme​ ​(DNA​ ​polymerase)​ ​and​ ​Mg​2+
○ dNTPs​ ​(dATP,​ ​dCTP,​ ​dGTP,​ ​dTTP),​ ​provide​ ​energy​ ​for​ ​polymerisation
○ Single-stranded​ ​template​ ​DNA
○ 3’-OH​ ​primer
● Process:
○ Initial​ ​denaturation​ ​of​ ​DNA​ ​at​ ​95​o​C​ ​for​ ​5​ ​minutes
■ Separates​ ​double-stranded​ ​DNA​ ​into​ ​single​ ​strands​ ​and​ ​detaches​ ​any
existing​ ​primers​ ​bound​ ​to​ ​the​ ​DNA
○ Primer​ ​annealing​ ​stage​ ​at​ ​55​o​C​ ​for​ ​1​ ​minute
○ Extension​ ​stage​ ​at​ ​72​o​C​ ​for​ ​1​ ​minute
■ Creation​ ​of​ ​new​ ​DNA​ ​complementary​ ​strands
○ Steps​ ​3-4​ ​are​ ​repeated​ ​for​ ​34​ ​cycles
○ Final​ ​extension​ ​stage​ ​at​ ​72​o​C​ ​for​ ​10​ ​minutes
○ Gel​ ​electrophoresis​ ​is​ ​used​ ​to​ ​separate​ ​the​ ​amplified​ ​DNA​ ​into​ ​size​ ​bands
--------------------------------------------------------------------------------------------------------------------------5:​ ​THE​ ​EUKARYOTIC​ ​CELL​ ​CYCLE
● S​ ​phase​ ​=​ ​synthesis​ ​(DNA​ ​replication)
● M​ ​phase​ ​=​ ​mitosis
● G1​ ​and​ ​G2​ ​=​ ​growth​ ​phases
Mitosis
● Prophase​ ​(early)
○ Chromosomes​ ​start​ ​to​ ​condense​ ​into​ ​chromatin
○ Mitotic​ ​spindle​ ​begins​ ​to​ ​form​ ​at​ ​the​ ​centromeres
○ The​ ​nucleolus​ ​disappears
● Prophase​ ​(late)
○ Chromosomes​ ​finish​ ​condensing
○ The​ ​nuclear​ ​envelope​ ​breaks​ ​down,​ ​releasing​ ​the​ ​chromosomes
○ More​ ​mitotic​ ​spindle​ ​forms,​ ​and​ ​some​ ​of​ ​the​ ​microtubules​ ​start​ ​to​ ​“capture”
chromosomes
● Metaphase
○ Chromosomes​ ​align​ ​along​ ​the​ ​equator
● Anaphase
Joanna​ ​Griffith​ ​(2017)

○ The​ ​spindle​ ​fibres​ ​pull​ ​the​ ​sister​ ​chromatids​ ​apart
● Telophase
○ Mitotic​ ​spindle​ ​is​ ​broken​ ​down
○ Two​ ​new​ ​nuclei​ ​form,​ ​along​ ​with​ ​nuclear​ ​membranes​ ​and​ ​nucleoli
○ Chromosomes​ ​decondense
● Cytokinesis
○ Division​ ​of​ ​the​ ​cytoplasm​ ​to​ ​form​ ​two​ ​new​ ​daughter​ ​cells
--------------------------------------------------------------------------------------------------------------------------5&6:​ ​BACTERIAL​ ​GENETICS
Genetic​ ​material
● Single​ ​circular​ ​double-stranded​ ​DNA​ ​molecule​ ​(chromosome/nucleoid)
○ No​ ​histone​ ​proteins
○ Associated​ ​with​ ​Mg​2+​​ ​and​ ​polyamines
■ Spermine
■ Spermidine
■ Putrescine
● Bacterial​ ​cells​ ​may​ ​also​ ​contain​ ​plasmids​ ​(smaller​ ​circles​ ​of​ ​DNA)
○ Can​ ​be​ ​passed​ ​between​ ​cells​ ​by​ ​conjugation
Replication
● Prokaryotes​ ​only​ ​have​ ​one​ ​origin​ ​of​ ​replication
● Lagging​ ​strand​ ​synthesis:
○ Primase​ ​synthesises​ ​short​ ​RNA​ ​oligonucleotides​ ​(primers)​ ​copied​ ​from​ ​DNA
○ DNA​ ​polymerase​ ​III​ ​elongates​ ​RNA​ ​primers​ ​with​ ​new​ ​DNA
○ DNA​ ​polymerase​ ​I​ ​removes​ ​RNA​ ​at​ ​the​ ​5’​ ​end​ ​of​ ​the​ ​neighbouring​ ​fragment
and​ ​fills​ ​the​ ​gap
○ DNA​ ​ligase​ ​connects​ ​adjacent​ ​fragments
Gene​ ​expression
● Prokaryotes​ ​do​ ​not​ ​have​ ​a​ ​nuclear​ ​membrane​ ​and​ ​only​ ​have​ ​one​ ​cytoplasmic
compartment,​ ​so​ ​they​ ​undergo​ ​coupled​ ​transcription​ ​and​ ​translation
○ Transcription​ ​and​ ​translation​ ​occur​ ​simultaneously​ ​in​ ​the​ ​cytoplasm
● Genes​ ​of​ ​related​ ​function​ ​are​ ​often​ ​clustered​ ​into​ ​operons
○ An​ ​operon​ ​has​ ​one​ ​promoter,​ ​and​ ​all​ ​genes​ ​in​ ​an​ ​operon​ ​are​ ​transcribed
together
--------------------------------------------------------------------------------------------------------------------------7:​ ​BIOTECHNOLOGY
Gene​ ​sequencing
● Genomics:​ ​everything​ ​from​ ​sequencing​ ​genomes,​ ​ascribing​ ​functions​ ​to​ ​genes,​ ​and
studying​ ​the​ ​structure​ ​of​ ​genes​ ​(gene​ ​architecture)
○ By​ ​studying​ ​an​ ​individual’s​ ​entire​ ​genome,​ ​we​ ​can​ ​see​ ​which​ ​genes​ ​are​ ​active
at​ ​particular​ ​times​ ​and​ ​under​ ​different​ ​environmental​ ​conditions,​ ​and​ ​see​ ​how
these​ ​affect​ ​outward​ ​characteristics
● Genome:​ ​all​ ​of​ ​the​ ​genes​ ​contained​ ​within​ ​an​ ​organism
● DNA​ ​sequencing
Joanna​ ​Griffith​ ​(2017)

Techniques​ ​used​ ​to​ ​produce​ ​millions​ ​of​ ​copies​ ​of​ ​short​ ​pieces​ ​of​ ​DNA
■ Nucleotides​ ​have​ ​fluorescent​ ​dyes​ ​attached,​ ​which​ ​are​ ​detected​ ​by
the​ ​sequencer
○ The​ ​ultimate​ ​goal​ ​is​ ​to​ ​determine​ ​the​ ​order​ ​of​ ​nucleotides​ ​throughout​ ​all​ ​of​ ​the
genetic​ ​material​ ​of​ ​an​ ​organism
○ Whole​ ​genome​ ​shotgun​ ​sequencing​ ​(allows​ ​entire​ ​genomes​ ​to​ ​be​ ​sequenced)
■ Breaks​ ​the​ ​genomic​ ​DNA​ ​into​ ​many​ ​small​ ​fragments
● Fragments​ ​are​ ​cloned​ ​into​ ​vectors
● The​ ​collection​ ​of​ ​fragments​ ​is​ ​called​ ​a​ ​genomic​ ​library
● Fragments​ ​can​ ​then​ ​be​ ​sequenced​ ​using​ ​a​ ​process​ ​called
chain​ ​termination​ ​DNA​ ​sequencing
○ Sanger​ ​sequencing
■ Depends​ ​on​ ​DNA​ ​replication​ ​machinery
■ Requires​ ​a​ ​3’​ ​OH​ ​group
○ Cycle​ ​sequencing
○ Chain​ ​termination​ ​sequencing
○ Whole​ ​genome​ ​shotgun​ ​sequencing
● How​ ​do​ ​we​ ​deal​ ​with​ ​genomic​ ​data?
○ Bioinformatics:​ ​the​ ​application​ ​of​ ​information​ ​technology​ ​to​ ​molecular​ ​biology
■ Primary​ ​goal​ ​is​ ​to​ ​increase​ ​our​ ​understanding​ ​of​ ​biological​ ​processes
■ Focus​ ​on​ ​developing​ ​and​ ​applying​ ​computationally​ ​intensive
techniques
● What​ ​sorts​ ​of​ ​questions​ ​can​ ​we​ ​ask​ ​of​ ​collected​ ​data?
○ Sequence​ ​alignment
○ Gene​ ​discovery
○ Gene​ ​assembly
○ Protein​ ​structure​ ​prediction
○ Prediction​ ​of​ ​gene​ ​expression
○ Modelling​ ​evolutionary​ ​relationships​ ​(eg
...
​ ​BamHI​ ​is​ ​from​ ​Bacillus​ ​amyloliquefaciens
● Recognises​ ​and​ ​cuts​ ​specific​ ​gene​ ​sequence
○

Joanna​ ​Griffith​ ​(2017)

○

●

●

●

●

This​ ​sequence​ ​occurs​ ​every​ ​4096​ ​base​ ​pairs
(therefore,​ ​an​ ​E
...
​ ​cloning​ ​DNA-recombinant​ ​E
...
​ ​DNA/RNA​ ​hybridisation
(measuring​ ​the​ ​degree​ ​of​ ​genetic​ ​similarity​ ​between​ ​pools​ ​of​ ​DNA
sequences)
■ Can​ ​be​ ​used​ ​to​ ​search​ ​for​ ​an​ ​expressed​ ​protein​ ​with​ ​an​ ​antibody
The​ ​power​ ​of​ ​recombinant​ ​DNA​ ​technology
○ Can​ ​clone​ ​genes​ ​from​ ​any​ ​organism
○ Can​ ​study​ ​any​ ​individual​ ​gene
■ Eg
...
​ ​microarrays)
○ Used​ ​in​ ​reverse​ ​genetics
■ The​ ​role​ ​of​ ​genes​ ​is​ ​investigated​ ​by​ ​measuring​ ​its​ ​effect​ ​on​ ​phenotype
○ Used​ ​in​ ​gene​ ​targeting
■ In​ ​vivo​ ​gene​ ​disruptions​ ​or​ ​deletions
○ Used​ ​in​ ​in​ ​vitro​ ​mutagenesis
■ Gene​ ​tagging​ ​(adding​ ​GFP​ ​(green​ ​fluorescent​ ​protein)​ ​or​ ​RFP​ ​(red
fluorescent​ ​protein))
■ Can​ ​reintroduce​ ​mutated​ ​genes​ ​in​ ​many​ ​cases​ ​(eg
...
​ ​Hepatitis​ ​B
--------------------------------------------------------------------------------------------------------------------------●

7:​ ​PCR​ ​AND​ ​GEL​ ​ELECTROPHORESIS
Polymerase​ ​Chain​ ​Reaction​ ​(PCR)
● Individual​ ​gene​ ​replication​ ​without​ ​cloning
● In​ ​vitro​ ​DNA​ ​synthesis​ ​reaction
○ Uses​ ​DNA​ ​+​ ​DNA​ ​polymerase​ ​+​ ​primers
○ Repeated​ ​many​ ​times,​ ​“chain​ ​reaction”
● Process:
○ Initial​ ​denaturation​ ​of​ ​DNA​ ​at​ ​95​°C​ ​for​ ​5​ ​minutes
■ Separates​ ​double-stranded​ ​DNA​ ​into​ ​single​ ​strands​ ​and​ ​detaches​ ​any
existing​ ​primers​ ​bound​ ​to​ ​the​ ​DNA
○ Primer​ ​annealing​ ​stage​ ​at​ ​55°C​ ​for​ ​1​ ​minute
○ Extension​ ​stage​ ​at​ ​72°C​ ​for​ ​1​ ​minute
■ Creation​ ​of​ ​new​ ​DNA​ ​complementary​ ​strands
○ Steps​ ​1-3​ ​are​ ​repeated​ ​for​ ​34​ ​cycles
○ Final​ ​extension​ ​stage​ ​at​ ​72°C​ ​for​ ​10​ ​minutes
○ Gel​ ​electrophoresis​ ​is​ ​used​ ​to​ ​separate​ ​the​ ​amplified​ ​DNA​ ​into​ ​size​ ​bands
● 30​ ​cycles​ ​of​ ​PCR​ ​produce​ ​about​ ​a​ ​10​6​-fold​ ​amplification
○ 1pg​ ​to​ ​1µg​ ​of​ ​DNA,​ ​enough​ ​to​ ​analyse​ ​on​ ​gel
● Only​ ​the​ ​DNA​ ​between​ ​primers​ ​is​ ​amplified
○ Specific​ ​sequences​ ​can​ ​be​ ​amplified​ ​from​ ​a​ ​complex​ ​mixture​ ​of​ ​DNA
○ The​ ​ends​ ​of​ ​the​ ​amplified​ ​fragment​ ​are​ ​defined​ ​by​ ​two​ ​primers
● Very​ ​powerful​ ​tool,​ ​with​ ​many​ ​research​ ​and​ ​applied​ ​uses
○ Detection​ ​of​ ​pathogens​ ​in​ ​water/blood
○ Genetic​ ​fingerprinting
○ Forensic​ ​analysis
Joanna​ ​Griffith​ ​(2017)

○ Diagnosis​ ​of​ ​genetic​ ​disorders
○ Prenatal​ ​diagnosis
○ Analysis​ ​of​ ​ancient​ ​DNA
○ Detection​ ​of​ ​insecticide​ ​resistance
● Limitations
○ Sequence​ ​information​ ​is​ ​required​ ​in​ ​order​ ​to​ ​design​ ​the​ ​primers
○ Limit​ ​on​ ​length​ ​of​ ​amplified​ ​fragments
○ Significant​ ​in​ ​vitro​ ​mutation​ ​rate
○ Very​ ​sensitive​ ​to​ ​exact​ ​reaction​ ​conditions
--------------------------------------------------------------------------------------------------------------------------8:​ ​REVISION
--------------------------------------------------------------------------------------------------------------------------9:​ ​PATTERNS​ ​AND​ ​PRINCIPLES​ ​OF​ ​HEREDITY:​ ​HOW​ ​ARE​ ​TRAITS​ ​TRANSMITTED?
The​ ​particulate​ ​theory​ ​of​ ​inheritance
● Characters​ ​are​ ​distinct​ ​and​ ​hereditary​ ​determinants​ ​(genes)​ ​are​ ​particulate​ ​in​ ​nature
● Each​ ​adult​ ​has​ ​two​ ​genes​ ​for​ ​each​ ​character
○ Different​ ​forms​ ​of​ ​the​ ​genes​ ​are​ ​called​ ​alleles
● Members​ ​of​ ​the​ ​gene​ ​pair​ ​segregate​ ​equally​ ​into​ ​gametes
○ Different​ ​genes​ ​assort​ ​independently​ ​in​ ​gametes
● Fusion​ ​of​ ​gametes​ ​at​ ​fertilisation​ ​restores​ ​the​ ​pair​ ​of​ ​genes​ ​and​ ​is​ ​random
Monohybrid​ ​crosses​ ​and​ ​law​ ​of​ ​segregation
● Eg
...
​ ​YY)
● Heterozygote:​ ​different​ ​alleles​ ​at​ ​a​ ​locus​ ​(eg
...
​ ​seed​ ​colour:​ ​Y​ ​=​ ​yellow,​ ​y​ ​=​ ​green​ ​and​ ​seed​ ​shape:​ ​R​ ​=​ ​round,​ ​r​ ​=​ ​wrinkly
■ Formation​ ​of​ ​gametes
● Rr​ ​x​ ​Yy
○ RY​ ​=​ ​¼
○ Ry​ ​=​ ​¼
○ rY​ ​=​ ​¼
○ ry​ ​=​ ​¼
■ The​ ​two​ ​traits​ ​are​ ​independent
● The​ ​9​ ​:​ ​3​ ​:​ ​3​ ​:​ ​1​ ​ratio​ ​is​ ​a​ ​random​ ​combination​ ​of​ ​two
independent​ ​3​ ​:​ ​1​ ​ratios
--------------------------------------------------------------------------------------------------------------------------●
●

10:​ ​RNA​ ​INTERFERENCE​ ​(RNAi):​ ​A​ ​MECHANISM​ ​FOR​ ​SILENCING​ ​GENE
EXPRESSION
Major​ ​scientific​ ​discoveries​ ​often​ ​begin​ ​with​ ​a​ ​surprising​ ​(unexpected)​ ​result
● Control​ ​of​ ​gene​ ​expression
○ Around​ ​1990,​ ​molecular​ ​biologists​ ​obtained​ ​a​ ​number​ ​of​ ​unexpected​ ​results
that​ ​were​ ​difficult​ ​to​ ​explain,​ ​given​ ​what​ ​was​ ​understood​ ​about​ ​control​ ​of​ ​gene
expression​ ​at​ ​the​ ​time
■ The​ ​most​ ​striking​ ​results​ ​were​ ​observed​ ​by​ ​plant​ ​biologists​ ​trying​ ​to
increase​ ​colour​ ​intensity​ ​of​ ​petunias​ ​by​ ​introducing​ ​a​ ​gene​ ​that​ ​leads
to​ ​the​ ​formation​ ​of​ ​red​ ​pigment
● Hypothesis:​ ​extra​ ​copies​ ​of​ ​a​ ​gene​ ​will​ ​result​ ​in​ ​more​ ​pigment,
leading​ ​to​ ​more​ ​intensely-coloured​ ​flowers
○ Actually​ ​found​ ​that​ ​flowers​ ​got​ ​less​ ​pigmented
■ How​ ​did​ ​adding​ ​extra​ ​copies​ ​of​ ​a​ ​gene​ ​that
enhances​ ​pigment​ ​production​ ​result​ ​in​ ​less
pigment?
● Control​ ​injection​ ​flowers​ ​had​ ​more
mRNA​ ​than​ ​the​ ​experimental​ ​injection
flowers
○ Reduced​ ​mRNA​ ​will​ ​lead​ ​to​ ​less
enzyme​ ​and​ ​therefore​ ​less
pigment​ ​production,​ ​so​ ​there​ ​will
be​ ​less​ ​pigment,​ ​but​ ​how​ ​does
increasing​ ​the​ ​number​ ​of​ ​copies
of​ ​the​ ​gene​ ​lead​ ​to​ ​reduced
mRNA?
dsRNA​ ​reduces​ ​levels​ ​of​ ​mRNA​ ​with​ ​matching​ ​nucleotide​ ​sequences,​ ​resulting​ ​in
gene​ ​silencing
● Andrew​ ​Fire​ ​and​ ​Craig​ ​Mello​ ​(1998)
○ Working​ ​on​ ​gene​ ​expression​ ​in​ ​the​ ​nematode​ ​C
...
​ ​p19)​ ​that
suppress​ ​gene​ ​silencing
○ microRNAs,​ ​which​ ​act​ ​like​ ​RNAi,​ ​have​ ​been​ ​shown​ ​to​ ​be​ ​key​ ​regulators​ ​in
many​ ​biological​ ​processes,​ ​such​ ​as​ ​development,​ ​cell​ ​birth​ ​and​ ​death,​ ​and
cancer
Is​ ​there​ ​clinical​ ​potential​ ​for​ ​RNAi?
● RNAi​ ​has​ ​the​ ​potential​ ​to​ ​specifically​ ​target​ ​gene​ ​expression,​ ​so​ ​has​ ​the​ ​potential​ ​to
fight​ ​almost​ ​every​ ​disease
● How​ ​close​ ​are​ ​we​ ​to​ ​seeing​ ​RNAi​ ​transform​ ​medicine?
○ Eg
...
​ ​hepatitis​ ​C
■ In​ ​2002,​ ​researchers​ ​at​ ​Stanford​ ​University​ ​announced​ ​that​ ​their​ ​RNAi
treatment​ ​had​ ​controlled​ ​the​ ​hepatitis​ ​C​ ​virus​ ​in​ ​lab​ ​mice
○ Eg
...
​ ​genetic​ ​contraceptives​ ​(not​ ​hormonal)
○ Currently,​ ​stem​ ​cells​ ​seem​ ​to​ ​be​ ​more​ ​useful​ ​than​ ​RNAi
--------------------------------------------------------------------------------------------------------------------------11:​ ​PATTERNS​ ​AND​ ​PRINCIPLES​ ​OF​ ​HEREDITY:​ ​WHAT​ ​COMPLEXITIES​ ​CAN​ ​BE
ENCOUNTERED​ ​IN​ ​RELATING​ ​GENOTYPE​ ​TO​ ​PHENOTYPE?
Interactions​ ​between​ ​alleles​ ​of​ ​a​ ​gene
● Dominance​ ​is​ ​not​ ​always​ ​complete
○ Incomplete​ ​dominance:​ ​heterozygotes​ ​show​ ​an​ ​intermediate​ ​(eg
...
​ ​in​ ​butterflies,​ ​different​ ​colours​ ​are​ ​more​ ​or​ ​less​ ​present​ ​in​ ​wing​ ​colouration
due​ ​to​ ​differences​ ​in​ ​dominance​ ​of​ ​colouration​ ​genes
○ Co-dominance​ ​:​ ​heterozygotes​ ​show​ ​phenotypes​ ​of​ ​both​ ​alleles
○ Eg
...
​ ​agouti​ ​-​ ​‘a’​ ​black​ ​and​ ​‘at’
black/yellow)
○ Eg
...
​ ​the​ ​gene​ ​involved​ ​in​ ​cilia​ ​and​ ​flagella​ ​production,​ ​if​ ​mutant,​ ​causes
respiratory​ ​problems​ ​(failure​ ​to​ ​clear​ ​airways)​ ​and​ ​sterility​ ​(sperm​ ​don’t​ ​have
normal​ ​motility)
○ Lethal​ ​alleles​ ​can​ ​cause​ ​skewed​ ​phenotypic​ ​ratios​ ​(missing​ ​from​ ​progeny)​ ​as
carriers​ ​of​ ​these​ ​alleles​ ​do​ ​not​ ​survive​ ​to​ ​be​ ​born
○ Eg
...
​ ​coat​ ​colour​ ​in​ ​mammals
■ Determined​ ​by​ ​at​ ​least​ ​5​ ​major​ ​genes

Joanna​ ​Griffith​ ​(2017)

A​ ​gene:​ ​determines​ ​the​ ​distribution​ ​of​ ​pigment​ ​in​ ​the​ ​hair​ ​(A​ ​=
agouti,​ ​a​ ​=​ ​solid)
● B​ ​gene:​ ​determines​ ​the​ ​colour​ ​of​ ​pigment​ ​in​ ​the​ ​hair​ ​(B​ ​=
black,​ ​b​ ​=​ ​brown)
● C​ ​gene:​ ​permits​ ​colour​ ​expression​ ​(C​ ​=​ ​colour​ ​expressed,​ ​c​ ​=
no​ ​colour)
● D​ ​gene:​ ​controls​ ​intensity​ ​of​ ​pigment​ ​specific​ ​by​ ​other​ ​genes
(D​ ​=​ ​full​ ​expression,​ ​d​ ​=​ ​dilute)
● S​ ​gene:​ ​controls​ ​distribution​ ​of​ ​pigment​ ​(S​ ​=​ ​solid​ ​colour,​ ​s​ ​=
spotted​ ​(piebald))
● Alleles​ ​of​ ​one​ ​gene​ ​can​ ​mask​ ​the​ ​effects​ ​of​ ​alleles​ ​at​ ​another​ ​gene
○ Epistasis:​ ​a​ ​gene​ ​interaction​ ​in​ ​which​ ​the​ ​effects​ ​of​ ​an​ ​allele​ ​at​ ​one​ ​gene​ ​hide
the​ ​effects​ ​of​ ​alleles​ ​at​ ​another​ ​gene
● Eg
...
​ ​AaBb​ ​x​ ​AaBb
■ Gametes:
● AB​ ​=​ ​¼
● Ab​ ​=​ ​¼
● aB​ ​=​ ​¼
● ab​ ​=​ ​¼
● But​ ​during​ ​meiosis​ ​I,​ ​chromatids​ ​from​ ​homologous​ ​pairs​ ​can
exchange​ ​strands​ ​in​ ​a​ ​process​ ​known​ ​as​ ​recombination
○ If​ ​a​ ​geneticist​ ​were​ ​to​ ​closely​ ​examine​ ​the​ ​genetic​ ​makeup​ ​of​ ​a​ ​single,
autosomal​ ​chromosome​ ​from​ ​one​ ​of​ ​your​ ​cells,​ ​that​ ​chromosome​ ​would​ ​be
found​ ​to​ ​be​ ​a​ ​mosaic​ ​of​ ​genes​ ​derived​ ​from​ ​just​ ​two​ ​of​ ​your​ ​grand-parents​ ​either​ ​your​ ​maternal​ ​grandparents​ ​or​ ​your​ ​paternal​ ​grandparents
--------------------------------------------------------------------------------------------------------------------------13:​ ​VARIATION​ ​IN​ ​CHROMOSOME​ ​NUMBER​ ​AND​ ​STRUCTURE:​ ​LARGE-SCALE
CHROMOSOMAL​ ​CHANGES
Changes​ ​in​ ​chromosome​ ​number
● Organisms​ ​with​ ​multiples​ ​of​ ​the​ ​basic​ ​chromosome​ ​set​ ​(genome)​ ​are​ ​referred​ ​to​ ​as
euploid
○ Chromosome​ ​number​ ​can​ ​vary​ ​among​ ​closely​ ​related​ ​species
■ Eg
...

■ An​ ​individual​ ​of​ ​a​ ​typically​ ​diploid​ ​species​ ​that​ ​has​ ​only​ ​one​ ​set​ ​of
chromosomes​ ​is​ ​called​ ​a​ ​monoploid​ ​(rather​ ​than​ ​a​ ​haploid,​ ​which​ ​is
the​ ​normal​ ​condition​ ​for​ ​some​ ​species)
■ May​ ​result​ ​in​ ​abnormal​ ​development,​ ​but​ ​not​ ​always
● Eg
...
​ ​tetraploid​ ​Australian​ ​frogs​ ​and​ ​Pacific​ ​oysters

Joanna​ ​Griffith​ ​(2017)

Individuals​ ​whose​ ​chromosome​ ​number​ ​differs​ ​by​ ​one​ ​or​ ​a​ ​small​ ​number​ ​of
chromosomes​ ​are​ ​referred​ ​to​ ​as​ ​aneuploid
○ An​ ​aneuploid​ ​can​ ​have​ ​a​ ​chromosome​ ​number​ ​either​ ​greater​ ​or​ ​smaller​ ​than
that​ ​of​ ​the​ ​wild​ ​type
■ For​ ​autosomes​ ​in​ ​a​ ​diploid​ ​organism:
● The​ ​aneuploid​ ​2n+1​ ​is​ ​trisomic​ ​(“three-bodies”)
● The​ ​aneuploid​ ​2n-1​ ​is​ ​monosomic
● The​ ​aneuploid​ ​2n-2​ ​is​ ​nullisomic
■ For​ ​the​ ​sex​ ​chromosomes,​ ​the​ ​notation​ ​lists​ ​the​ ​copies​ ​of​ ​each
chromosome​ ​(eg
...
​ ​trisomy​ ​21​ ​(causes​ ​by​ ​non-disjunction​ ​in​ ​the​ ​mother’s
egg​ ​in​ ​90%​ ​of​ ​cases)​ ​causes​ ​Down​ ​Syndrome
■ Klinefelter​ ​syndrome​ ​results​ ​from​ ​an​ ​XXY​ ​karyotype​ ​(the​ ​extra
chromosome​ ​can​ ​come​ ​from​ ​both​ ​the​ ​mother​ ​or​ ​the​ ​father​ ​in​ ​this​ ​case)
○ Why​ ​are​ ​aneuploids​ ​so​ ​much​ ​more​ ​abnormal​ ​than​ ​polyploids?
○ Why​ ​does​ ​aneuploidy​ ​for​ ​each​ ​chromosome​ ​have​ ​its​ ​own​ ​characteristic
phenotypic​ ​effects?
○ Why​ ​are​ ​monosomics​ ​typically​ ​more​ ​severe​ ​than​ ​the​ ​corresponding
trisomics?
○ Gene​ ​balance:​ ​genes​ ​have​ ​evolved​ ​to​ ​function​ ​in​ ​a​ ​diploid​ ​genetic
background,​ ​and​ ​disrupting​ ​that​ ​background​ ​disrupts​ ​their​ ​function
■ Haplo-abnormal​ ​genes
■ Triplo-abnormal​ ​genes
■ Expression​ ​of​ ​deleterious​ ​alleles​ ​on​ ​monosomic​ ​autosomes​ ​(in​ ​the
absence​ ​of​ ​wild-type​ ​counterpart)
■ Having​ ​only​ ​one​ ​gene​ ​copy​ ​(monosomic)​ ​is​ ​worse​ ​than​ ​having​ ​three
(trisomic)
Changes​ ​in​ ​chromosome​ ​structure
● Chromosomes​ ​can​ ​have​ ​missing​ ​pieces:​ ​deletions
○ A​ ​deletion​ ​is​ ​the​ ​loss​ ​of​ ​part​ ​of​ ​one​ ​chromosome​ ​arm
○ Deletions​ ​can​ ​be​ ​small,​ ​only​ ​covering​ ​a​ ​part​ ​of​ ​one​ ​gene,​ ​or​ ​large,​ ​with
chromosomes​ ​missing​ ​pieces​ ​large​ ​enough​ ​to​ ​be​ ​visualised​ ​on​ ​a​ ​karyotype
●

Joanna​ ​Griffith​ ​(2017)

Eg
...
3​ ​=​ ​cat-like​ ​cry
○ Sp15
...
​ ​Williams​ ​syndrome
● FISH​ ​allows​ ​identification​ ​of​ ​the​ ​deletion​ ​of​ ​one​ ​elastin​ ​gene
● More​ ​than​ ​25​ ​genes​ ​deleted​ ​on​ ​chromosome​ ​7
○ ELN​ ​(elastin)​ ​gene​ ​=​ ​connective​ ​tissue​ ​abnormalities
and​ ​cardiovascular​ ​disease
○ CLIP2,​ ​GTF21,​ ​GTF21RD1,​ ​LIMK1​ ​=​ ​problems​ ​with
visual​ ​spatial​ ​tasks
○ NCF1​ ​(neutrophil​ ​cytosolic​ ​factor​ ​1)​ ​=​ ​related​ ​to​ ​risk​ ​of
developing​ ​hypertension​ ​if​ ​NOT​ ​deleted​ ​(part​ ​of
NADPH​ ​oxidase​ ​which​ ​increases​ ​reactive​ ​oxygen
species​ ​and​ ​blood​ ​vessel​ ​changes)
○ Most​ ​deletions​ ​are​ ​not​ ​inherited​ ​(no​ ​family​ ​history)
■ Usually​ ​a​ ​result​ ​of​ ​random​ ​events​ ​during​ ​the
production​ ​of​ ​eggs​ ​and​ ​sperm
Chromosomes​ ​can​ ​have​ ​extra​ ​pieces:​ ​duplications
○ Duplications​ ​play​ ​an​ ​important​ ​role​ ​in​ ​evolution​ ​of​ ​the​ ​genome​ ​(eg
...
​ ​normal​ ​eye​ ​colour​ ​is​ ​red,​ ​but​ ​Morgan​ ​isolated​ ​a​ ​mutant
with​ ​white​ ​eyes​ ​(w​ ​gene)
Joanna​ ​Griffith​ ​(2017)

Cross​ ​1:
■ White​ ​male​ ​x​ ​red​ ​female​ ​=​ ​F​1​​ ​all​ ​have​ ​red​ ​eyes​ ​(red​ ​is​ ​dominant)
○ Cross​ ​2:
■ F​1​​ ​female​ ​x​ ​F​1​​ ​male​ ​(both​ ​red)​ ​=​ ​F​2​​ ​3:1​ ​red:white,​ ​but​ ​all​ ​white-eyed
flies​ ​are​ ​male,​ ​and​ ​there​ ​is​ ​a​ ​2:1​ ​ratio​ ​of​ ​females:males​ ​among​ ​the
red-eyed​ ​flies
○ Cross​ ​3​ ​(back​ ​cross):
■ White​ ​male​ ​x​ ​F​1​​ ​female​ ​(red)​ ​=​ ​ratios​ ​of​ ​1:1​ ​(red​ ​males:red​ ​females)
and​ ​1:1​ ​(white​ ​males:white​ ​females)
○ Cross​ ​4​ ​(reciprocal​ ​cross):
■ White​ ​female​ ​x​ ​F1​​ ​ ​male​ ​(red)​ ​=​ ​white​ ​males​ ​and​ ​red​ ​females
○ Cytological​ ​data:
■ Drosophila​ ​has​ ​4​ ​pairs​ ​of​ ​chromosomes,​ ​3​ ​pairs​ ​are​ ​homomorphic​ ​(the
same)​ ​but​ ​1​ ​pair​ ​is​ ​heteromorphic
■ Females​ ​are​ ​XX,​ ​males​ ​are​ ​XY
■ What​ ​other​ ​data​ ​could​ ​give​ ​clues​ ​to​ ​the​ ​inheritance​ ​pattern​ ​of​ ​eye
colour
○ Hypothesis:​ ​the​ ​gene​ ​encoding​ ​eye​ ​colour​ ​is​ ​on​ ​the​ ​X​ ​chromosome
■ Cross​ ​1:
● XWXW​ ​x​ ​XwY​ ​(red​ ​female​ ​x​ ​white​ ​male)
○ Produces​ ​red​ ​females​ ​and​ ​red​ ​males​ ​(all​ ​red​ ​offspring)
■ Cross​ ​2:
● XWXw​ ​x​ ​XWY​ ​(F​1​​ ​female,​ ​red​ ​x​ ​F1​​ ​ ​male,​ ​red)
○ 3:1​ ​ratio​ ​of​ ​red:white,​ ​all​ ​white-eyed​ ​flies​ ​are​ ​male
● What​ ​about​ ​chickens​ ​and​ ​moths
○ Moths​ ​are​ ​different​ ​again,​ ​but​ ​the​ ​same​ ​basic​ ​rules​ ​still​ ​apply
○ Homomorphic​ ​sex​ ​(sex​ ​chromosomes​ ​are​ ​the​ ​same,​ ​XX)
○ Heteromorphic​ ​sex​ ​(sex​ ​chromosomes​ ​are​ ​different,​ ​XY)
○ The​ ​ZW​ ​system
■ Males​ ​are​ ​ZZ,​ ​females​ ​are​ ​ZW
■ Eg
...
​ ​some​ ​X-linked​ ​traits​ ​in​ ​humans
■ Red-green​ ​colorblindness
■ Haemophilia
■ Duchenne’s​ ​muscular​ ​dystrophy
○

Joanna​ ​Griffith​ ​(2017)

Males​ ​are​ ​hemizygous​ ​for​ ​genes​ ​on​ ​the​ ​X​ ​chromosome​ ​(effectively​ ​dominant​ ​as​ ​a
single​ ​copy)
X-linked​ ​recessive​ ​traits​ ​can​ ​be​ ​deduced​ ​from​ ​certain​ ​clues
● More​ ​males​ ​than​ ​females​ ​express​ ​the​ ​trait
● For​ ​a​ ​female​ ​to​ ​express​ ​the​ ​trait,​ ​the​ ​male​ ​parent​ ​must​ ​express​ ​it​ ​and​ ​the​ ​female
parent​ ​must​ ​either​ ​express​ ​it​ ​or​ ​be​ ​a​ ​carrier
● The​ ​characteristic​ ​often​ ​skips​ ​a​ ​generation
● If​ ​a​ ​female​ ​expresses​ ​the​ ​characteristic,​ ​all​ ​of​ ​her​ ​male​ ​offspring​ ​will​ ​express​ ​the​ ​trait
In​ ​female​ ​mammals,​ ​one​ ​X​ ​chromosome​ ​is​ ​inactivated​ ​early​ ​in​ ​development
● Inactivated​ ​X​ ​chromosomes​ ​can​ ​be​ ​seen​ ​as​ ​highly​ ​condensed​ ​‘Barr’​ ​bodies
● Inactivation​ ​is​ ​random​ ​-​ ​the​ ​maternal​ ​OR​ ​paternal​ ​X​ ​chromosome​ ​can​ ​be​ ​inactivated
(although​ ​there​ ​are​ ​some​ ​exceptions)
● Therefore,​ ​the​ ​female​ ​body​ ​is​ ​mosaic​ ​for​ ​genes​ ​on​ ​the​ ​X​ ​chromosomes
○ Eg
...
​ ​pr​ ​=​ ​purple​ ​eye,​ ​pr​+​​ ​=​ ​wild​ ​type​ ​(red​ ​eye)
● Eg
...
​ ​trans​ ​(different​ ​chromosome)
configuration​ ​of​ ​genes
○ Cis​ ​configuration:
■ Pr​+​pr​+​vg​+​vg​+​​ ​x​ ​prprvgvg
● F​1​​ ​pr​+​prvg​+​vg​ ​x​ ​prprvgvg
○ F​2​​ ​pr​+​vg​+​​ ​=​ ​1000,​ ​prvg​ ​=​ ​1000,
prvg​+​​ ​=​ ​150,​ ​pr​+​vg​ ​=​ ​150
○ Trans​ ​configuration:
■ Pr​+​pr​+​vgvg​ ​x​ ​prprvg​+​vg​+
● F​1​​ ​pr​+​prvg​+​vg​ ​x​ ​prprvgvg
○ F​2​​ ​pr​+​vg​+​​ ​=​ ​150,​ ​prvg​ ​=​ ​150,​ ​prvg​+
=​ ​1000,​ ​pr​+​vg​ ​=​ ​1000
● Non-parental​ ​genotypes​ ​arise​ ​due​ ​to​ ​‘crossing​ ​over’​ ​during​ ​meiosis​ ​I,​ ​and​ ​are​ ​called
recombinants
○ No​ ​crossover​ ​between​ ​genes​ ​results​ ​in​ ​100%​ ​parental​ ​genotypes
○ Crossover​ ​between​ ​genes​ ​results​ ​in​ ​some​ ​recombinant​ ​genotypes​ ​and​ ​some
parental​ ​genotypes
● The​ ​frequency​ ​of​ ​crossing​ ​over​ ​between​ ​two​ ​genes​ ​gives​ ​us​ ​information​ ​about​ ​the
physical​ ​distance​ ​between​ ​the​ ​genes​ ​on​ ​the​ ​chromosome
○ We​ ​can​ ​use​ ​recombination​ ​to​ ​produce​ ​a​ ​‘linkage​ ​map’​ ​of​ ​a​ ​chromosome
because​ ​the​ ​frequency​ ​of​ ​recombination​ ​between​ ​two​ ​genes​ ​will​ ​depend​ ​on
the​ ​distance​ ​between​ ​them
■ Bear​ ​in​ ​mind​ ​that​ ​the​ ​frequency​ ​of​ ​crossing​ ​over​ ​is​ ​not​ ​really​ ​linear
along​ ​the​ ​chromosome,​ ​and​ ​there​ ​are​ ​actually​ ​‘hot​ ​spots’​ ​for
recombination
○ One​ ​genetic​ ​map​ ​unit​ ​=​ ​the​ ​distance​ ​between​ ​genes​ ​for​ ​which​ ​one​ ​product​ ​of
meiosis​ ​is​ ​recombinant​ ​=​ ​centimorgan​ ​(100​ ​units/morgans)
○ Genetic​ ​maps​ ​are​ ​linear​ ​and​ ​additive​ ​(map​ ​units)
● Chromosomes​ ​can​ ​be​ ​‘mapped’​ ​using​ ​three-point​ ​crosses
○ Look​ ​at​ ​patterns​ ​-​ ​are​ ​there​ ​two​ ​or​ ​three​ ​classes​ ​of​ ​offspring
(common/uncommon/rare)
■ If​ ​there​ ​are​ ​three​ ​classes,​ ​you​ ​are​ ​likely​ ​dealing​ ​with​ ​three​ ​linked
genes
○ Determine​ ​parental​ ​and​ ​double​ ​recombinant​ ​types
○ Deduce​ ​gene​ ​order
○ Calculate​ ​distances​ ​between​ ​pairs​ ​of​ ​genes
○ Gene​ ​order​ ​can​ ​be​ ​deduced​ ​by​ ​comparing​ ​the​ ​parental​ ​genotypes​ ​to​ ​the
genotypes​ ​of​ ​the​ ​double​ ​recombinant​ ​offspring
--------------------------------------------------------------------------------------------------------------------------■

Joanna​ ​Griffith​ ​(2017)

16:​ ​SPECIAL​ ​ISSUES​ ​IN​ ​HUMAN​ ​GENETICS
Why​ ​do​ ​we​ ​need​ ​to​ ​consider​ ​human​ ​genetics​ ​separately?
○ Can’t​ ​do​ ​controlled​ ​crosses
○ Limited​ ​numbers​ ​of​ ​offspring
○ Human​ ​genetics​ ​is​ ​of​ ​medical​ ​importance
○ We​ ​are​ ​inherently​ ​interested​ ​in​ ​ourselves
● What​ ​sort​ ​of​ ​human​ ​traits​ ​are​ ​of​ ​genetic​ ​interest?
● How​ ​can​ ​we​ ​study​ ​human​ ​genetics?
Pedigree​ ​analysis
● Factors​ ​to​ ​consider​ ​in​ ​pedigrees:
○ Is​ ​the​ ​trait​ ​located​ ​on​ ​a​ ​sex​ ​chromosome​ ​or​ ​autosome?
■ Autosomal:​ ​not​ ​on​ ​a​ ​sex​ ​chromosome
■ Sex-linkage:​ ​located​ ​on​ ​one​ ​of​ ​the​ ​sex​ ​chromosomes
● Y-linked:​ ​only​ ​males​ ​carry​ ​the​ ​trait
● X-linked​ ​(recessive):​ ​sons​ ​inherit​ ​the​ ​disease​ ​from​ ​normal
parents
● Basic​ ​symbols:
○ A​ ​circle​ ​is​ ​female
○ A​ ​square​ ​is​ ​male
○ A​ ​horizontal​ ​line​ ​indicates​ ​a​ ​mating
○ Offspring​ ​are​ ​depicted​ ​below​ ​the​ ​parents
○ Filling​ ​in​ ​the​ ​symbol​ ​indicates​ ​that​ ​the​ ​trait​ ​is​ ​expressed
● Eg
...
​ ​albinism
○ Lack​ ​of​ ​pigmentation​ ​in​ ​eyes​ ​and​ ​skin
○ Expressed​ ​in​ ​both​ ​sexes​ ​at​ ​approximately​ ​equal​ ​frequency,​ ​so​ ​autosomal
○ Not​ ​expressed​ ​in​ ​every​ ​generation,​ ​so​ ​recessive
● Eg
...
​ ​trisomy​ ​21​ ​(Down​ ​Syndrome)
●

Joanna​ ​Griffith​ ​(2017)

○

○

● Tremendous​ ​variation​ ​in​ ​phenotype
■ Monosomy:​ ​only​ ​one​ ​chromosome​ ​present​ ​when​ ​there​ ​should​ ​be​ ​two
■ Trisomy:​ ​three​ ​chromosomes​ ​present​ ​when​ ​there​ ​should​ ​be​ ​two
■ Haplo-diploidy:​ ​all​ ​males​ ​are​ ​haploid
■ Most​ ​embryos​ ​with​ ​a​ ​trisomy​ ​or​ ​monosomy​ ​don’t​ ​develop
Genetic​ ​disorders​ ​can​ ​be​ ​caused​ ​by​ ​non-nuclear​ ​genes​ ​(mitochondrial)
■ Maternal​ ​(mitochondrial)​ ​inheritance:
● Inheritance​ ​through​ ​the​ ​maternal​ ​lineage
● Sperm​ ​do​ ​not​ ​contribute​ ​mitochondria​ ​to​ ​the​ ​embryo
■ Eg
...
​ ​phenylketonuria​ ​(PKU)
● Classic​ ​example​ ​of​ ​a​ ​single​ ​gene​ ​mutation​ ​in​ ​a​ ​biochemical
pathway​ ​leading​ ​to​ ​disease
● Biochemical​ ​defect​ ​detected​ ​in​ ​the​ ​1950s,​ ​neonatal​ ​screening
came​ ​into​ ​use​ ​in​ ​the​ ​1960s,​ ​allowing​ ​early​ ​diagnosis​ ​and
treatment
● PAH​ ​(phenylalanine​ ​hydroxylase)​ ​is​ ​used​ ​in​ ​the​ ​conversion​ ​of
ingested​ ​phenylalanine​ ​to​ ​tyrosine​ ​by​ ​the​ ​addition​ ​of​ ​an​ ​-OH
group​ ​to​ ​the​ ​phenyl​ ​ring
○ When​ ​PKU​ ​mutates,​ ​there​ ​is​ ​no​ ​active​ ​PAH​ ​so
phenylalanine​ ​builds​ ​up,​ ​producing​ ​toxic​ ​byproducts
that​ ​cause​ ​mental​ ​retardation
● Treatment:​ ​avoid​ ​dietary​ ​phenylalanine,​ ​thus​ ​reducing​ ​the
toxins​ ​and​ ​the​ ​mental​ ​retardation
○ Some​ ​patients​ ​did​ ​not​ ​respond​ ​to​ ​standard​ ​treatment
■ Mapped​ ​and​ ​cloned​ ​the​ ​PAH​ ​gene​ ​in​ ​1983,
discovered​ ​allelic​ ​heterogeneity​ ​in​ ​PKU​ ​as​ ​well
as​ ​the​ ​other​ ​loci​ ​involved​ ​(not​ ​a​ ​single​ ​gene
disorder)
● Genetic​ ​background​ ​is​ ​important
■ Eg
...
​ ​litter​ ​size,​ ​lifetime​ ​reproductive​ ​output,
longevity
● Variation​ ​in​ ​protein​ ​sequence
○ Allozyme​ ​electrophoresis
Joanna​ ​Griffith​ ​(2017)

Variation​ ​in​ ​DNA​ ​sequence
○ Microsatellites,​ ​DNA​ ​fingerprints,​ ​RFLPs,​ ​DNA
sequencing,​ ​etc
...
​ ​actual​ ​numbers​ ​of
offspring
Selection​ ​results​ ​in​ ​adaptation​ ​to​ ​the​ ​environment
■ A​ ​heritable​ ​variation​ ​in​ ​a​ ​trait​ ​(genetic​ ​diversity)​ ​is​ ​correlated​ ​with​ ​an
increase​ ​in​ ​reproductive​ ​success
● In​ ​response​ ​to​ ​this​ ​selection,​ ​alleles​ ​that​ ​confer​ ​advantages​ ​in
reproductive​ ​success​ ​increase​ ​in​ ​frequency​ ​in​ ​the​ ​population
■ A​ ​founder​ ​population​ ​separated​ ​into​ ​two​ ​environments​ ​will​ ​evolve
adaptations
● These​ ​adaptations​ ​are​ ​likely​ ​to​ ​be​ ​different​ ​between​ ​the​ ​two
populations​ ​if​ ​the​ ​environments​ ​are​ ​different​ ​from​ ​each​ ​other
Loss​ ​of​ ​genetic​ ​diversity​ ​reduces​ ​the​ ​ability​ ​of​ ​a​ ​population​ ​to​ ​adapt​ ​to
environmental​ ​changes
■ Eg
...
0
● Average​ ​heterozygosity​ ​=​ ​0
...
​ ​humans​ ​have​ ​>200​ ​alleles​ ​making
up​ ​the​ ​MHC,​ ​implying​ ​selective​ ​pressure​ ​to​ ​establish
and​ ​maintain​ ​polymorphism)
■ Reduction​ ​in​ ​MHC​ ​polymorphism​ ​would​ ​make​ ​a
population​ ​vulnerable​ ​to​ ​novel​ ​pathogens
○ In​ ​a​ ​cheetah​ ​colony​ ​in​ ​Oregon,​ ​in​ ​the​ ​USA,​ ​a​ ​feline
virus​ ​killed​ ​60%​ ​of​ ​the​ ​cheetahs​ ​but​ ​did​ ​not​ ​affect​ ​lions
in​ ​the​ ​same​ ​colony​ ​-​ ​is​ ​this​ ​a​ ​result​ ​of​ ​low​ ​MHC
diversity​ ​in​ ​cheetahs?
■ Merola​ ​(1994)
● Compared​ ​cheetah​ ​genetic​ ​variability
with​ ​other​ ​carnivorous​ ​vertebrates
●

○

○

Joanna​ ​Griffith​ ​(2017)

Of​ ​the​ ​24​ ​species​ ​surveyed,​ ​8
other​ ​species​ ​than​ ​cheetahs​ ​had
no​ ​heterozygosity
● Suggested​ ​that​ ​the​ ​Oregon​ ​virus​ ​was
only​ ​effective​ ​because​ ​of​ ​the​ ​high
density​ ​in​ ​captivity,​ ​not​ ​genetics
● Reviewed​ ​by​ ​May​ ​in​ ​1995,​ ​who​ ​found
evidence​ ​for​ ​genetic​ ​problems​ ​but​ ​noted
that​ ​environmental​ ​effects​ ​were​ ​also
important
Genetic​ ​adaptation​ ​to​ ​captivity​ ​and​ ​the​ ​ability​ ​to​ ​return​ ​to​ ​the​ ​wild
■ The​ ​IUCN​ ​has​ ​endorsed​ ​captive​ ​breeding​ ​programs
● Establish​ ​secure​ ​populations
● Educate​ ​and​ ​engage​ ​the​ ​public
● Research​ ​on​ ​the​ ​basic​ ​biology​ ​of​ ​a​ ​species
● Provide​ ​animals​ ​for​ ​reintroduction​ ​programs
■ Genetic​ ​adaptation​ ​to​ ​captivity​ ​is​ ​recognised​ ​as​ ​a​ ​serious​ ​threat​ ​to​ ​the
success​ ​of​ ​reintroductions
What​ ​happens​ ​when​ ​the​ ​assumption​ ​of​ ​infinite​ ​population​ ​size​ ​is​ ​violated?
■ Conservation​ ​biology​ ​is​ ​concerned​ ​with​ ​small​ ​and/or​ ​declining
populations​ ​where​ ​chance​ ​has​ ​a​ ​greater​ ​impact​ ​and​ ​selection​ ​is​ ​less
effective
■ Genetic​ ​drift​ ​-​ ​chance​ ​effects​ ​can​ ​override​ ​natural​ ​selection
● Small​ ​populations​ ​lead​ ​to​ ​sampling​ ​problems
○ Founder​ ​effect
■ The​ ​reduced​ ​genetic​ ​diversity​ ​of​ ​a​ ​population
when​ ​it​ ​is​ ​descended​ ​from​ ​a​ ​small​ ​number​ ​of
colonising​ ​ancestors
○ Bottleneck​ ​effect
■ A​ ​sharp​ ​reduction​ ​in​ ​the​ ​size​ ​of​ ​a​ ​population​ ​due
to​ ​environmental​ ​effects​ ​(eg
...
​ ​poaching)
● Genetic​ ​drift​ ​can​ ​lead​ ​to​ ​loss​ ​of​ ​alleles​ ​from​ ​the​ ​population​ ​or
fixation​ ​of​ ​a​ ​particular​ ​allele​ ​at​ ​a​ ​locus,​ ​thus​ ​allelic​ ​diversity​ ​is
lost​ ​at​ ​random
■ Inbreeding​ ​-​ ​a​ ​reduction​ ​in​ ​genetic​ ​diversity?
● In​ ​small​ ​populations,​ ​inbreeding​ ​is​ ​inevitable
● Recessive​ ​phenotypes​ ​are​ ​amplified​ ​in​ ​the​ ​population
● Inbreeding​ ​results​ ​in​ ​a​ ​loss​ ​of​ ​genetic​ ​diversity​ ​and​ ​exposes
rare​ ​deleterious​ ​alleles​ ​leading​ ​to​ ​reductions​ ​in​ ​reproductive
fitness​ ​(inbreeding​ ​depression)
○ Does​ ​inbreeding​ ​depression​ ​lead​ ​to​ ​reduced
reproduction​ ​and​ ​survival​ ​and​ ​increased​ ​risk​ ​of
extinction​ ​in​ ​wild​ ​populations?
■ Saccheri​ ​et​ ​al​ ​(1998)
○

○

○

Joanna​ ​Griffith​ ​(2017)

Genotyped​ ​female​ ​Granville​ ​fritillary
butterflies​ ​from​ ​42​ ​populations​ ​at​ ​8
polymorphic​ ​loci
○ Populations​ ​with​ ​less​ ​genetic
diversity​ ​are​ ​more​ ​likely​ ​to​ ​go
extinct
■ Genetic​ ​diversity
predicted​ ​extinction​ ​risk
after​ ​accounting​ ​for​ ​all
demographic,​ ​ecological,
and​ ​environmental
causes​ ​of​ ​extinction
■ Inbreeding​ ​reduced​ ​egg
hatch​ ​rate,​ ​larval​ ​survival,
and​ ​female​ ​lifespan​ ​(so
they​ ​had​ ​less​ ​time​ ​to​ ​lay
eggs)
○ What​ ​happens​ ​when​ ​the​ ​assumption​ ​of​ ​no​ ​migration​ ​between​ ​populations​ ​is
violated?
■ The​ ​introduction​ ​of​ ​immigrants​ ​from​ ​one​ ​population​ ​into​ ​another
reduces​ ​genetic​ ​differentiation​ ​among​ ​populations​ ​and​ ​may​ ​restore
genetic​ ​diversity
■ Migration​ ​can​ ​minimise​ ​inbreeding​ ​depression​ ​and​ ​the​ ​loss​ ​of​ ​genetic
diversity
● Delta​ ​Q​ ​=​ ​m(qm​ ​-​ ​qo)
● The​ ​change​ ​in​ ​the​ ​frequency​ ​of​ ​an​ ​allele​ ​depends​ ​on:
○ M​ ​=​ ​migration​ ​rate​ ​(the​ ​number​ ​of​ ​individuals​ ​migrating
in​ ​relative​ ​to​ ​the​ ​number​ ​in​ ​the​ ​original​ ​population)
○ Qm​ ​=​ ​the​ ​frequency​ ​of​ ​that​ ​allele​ ​in​ ​the​ ​migrant
population
○ Qo​ ​=​ ​the​ ​frequency​ ​of​ ​that​ ​allele​ ​in​ ​the​ ​original
population
■ Fragmented​ ​populations​ ​can​ ​be​ ​managed​ ​by​ ​providing​ ​“corridors”​ ​of
gene​ ​flow​ ​-​ ​is​ ​this​ ​always​ ​a​ ​good​ ​thing?
● Habitat​ ​loss​ ​has​ ​led​ ​to​ ​fragmented​ ​populations
● Populations​ ​show​ ​divergence​ ​in​ ​allele​ ​frequencies
● The​ ​smallest​ ​populations​ ​show​ ​the​ ​greatest​ ​loss​ ​of​ ​genetic
diversity
Genetics​ ​can​ ​resolve​ ​taxonomic​ ​uncertainties
● How​ ​do​ ​we​ ​define​ ​a​ ​species?
○ A​ ​group​ ​of​ ​closely​ ​related​ ​organisms​ ​that​ ​are​ ​reproductively​ ​isolated,​ ​and​ ​are
able​ ​to​ ​reproduce​ ​to​ ​produce​ ​fertile​ ​offspring
○ Tamarin​ ​species​ ​look​ ​the​ ​same​ ​and​ ​will​ ​interbreed​ ​if​ ​put​ ​together,​ ​but​ ​have
different​ ​ranges​ ​and​ ​significantly​ ​quantitative​ ​genetic​ ​differentiation
○ Why​ ​does​ ​it​ ​matter?
●

Joanna​ ​Griffith​ ​(2017)

The​ ​most​ ​serious​ ​weakness​ ​in​ ​sustaining​ ​current​ ​approaches​ ​to​ ​the
study​ ​of​ ​biological​ ​diversity​ ​is​ ​our​ ​limited​ ​ability​ ​to​ ​recognise
morphological​ ​variation
● Solution:​ ​microgenomics
Molecular​ ​genetics​ ​can​ ​be​ ​used​ ​in​ ​wildlife​ ​forensics
● Captive​ ​bred​ ​vs
...
​ ​exotic​ ​pet​ ​trade​ ​laundering​ ​of​ ​wild​ ​caught​ ​animals​ ​through​ ​captive
breeding​ ​centres​ ​(can​ ​be​ ​used​ ​to​ ​verify​ ​family​ ​relationships)
● Identification​ ​of​ ​species
○ Eg
...
​ ​identifying​ ​targeted​ ​elephant​ ​populations​ ​for​ ​ivory​ ​in​ ​order​ ​to​ ​pinpoint
anti-poaching​ ​schemes
● Illegal​ ​trade​ ​product​ ​identification
○ Eg

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