School of Biosciences
Genetics & Cell Biology (D211P1)
Dr Lee Garratt, genetics lecture summary
1
COMPLEMENTATION
1.
Chemical conversions in a biochemical pathway are carried out by enzymes
2.
Different enzymes are encoded by different genes
3.
Mutation of a gene may result in no active enzyme being formed and a block in the
biochemical pathway at a particular point
4.
DIPLOID organisms have two copies of each gene, HAPLOID organisms have only one.
5.
These copies are independent (usually) and if one is normal some active enzyme will be
formed - the mutant gene is said to be RECESSIVE.
6.
If a diploid has two defective copies of the same gene (double recessive), it will have a mutant
phenotype
7.
If the two defective genes in a diploid are different (and encode different enzyme steps) they
will COMPLEMENT each other. In other words, there will be a full set of normal genes, a full set
of active enzymes and a wild type phenotype.
8.
Two genes that complement each other are in different COMPLEMENTATION GROUPS. Those that
do not complement are in the same group.
9.
A similar effect is seen if two mutant haploids are grown close together, due to diffusion of
metabolic intermediates (CROSS FEEDING).
10.
A genetic unit defined by complementation is not necessarily a true gene and is called a
CISTRON.
COLLINEARITY OF GENE & PROTEIN
1.
Proteins are made up of chains of amino acids, each with particular chemical properties
2.
The nature and order of these may be vital for protein function
3.
Each mutation in a gene that destroys the function of a protein causes a specific change of one
amino acid for another
4.
Some mutations (SILENT MUTATIONS) do not affect protein function
5.
Each specific change of amino acid occurs at a specific place in the protein chain
6.
Mutations also occur at specific places in a gene (recombination map)
7.
The protein sequence and the genetic map are COLLINEAR
8.
Two different mutations that map close together but which can recombine may affect the same
amino acid. Therefore more than one mutable site (nucleotide) specifies each amino acid
Inherited human anaemias and thallasaemias
Haemoglobin
Subunit and amino acid
Change
Hb I
Hb G Honolulu
Hb Norfolk
Hb M Boston
Hb G Philadelphia





16
30
57
58
68
lys  asp
glu  gln
gly  asp
his  tyr
asn  lys
Hb S (sickle)
Hb C
Hb G San José
Hb E
Hb M Saskatoon
Hb Zürich
Hb M Milwaukee
Hb D  Punjab








6
6
7
26
63
63
67
125
glu  val
glu  lys
glu  gly
glu  lys
his  tyr
his  arg
val  glu
glu  gln
The tryptophan synthase (trpA) cistron
Positions of mutant loci on genetic map
446 487
223
23 187
58 169
212 215
234 235
Positions of altered amino acids in protein chain
175 177
183
The genetic map and the amino acid sequence are collinear (the changed amino acids and the mutant
loci appear in the same relative positions).
In E. coli, recombination can occur between mutant loci that are very close together (e.g. 58 and 169).
Several mutations have been found that affect the nature of the amino acid at position 212.
Genetic variant
Amino acid at position 212
Wild type
Mutant 23
Mutant 46
23  46 double mutant recombinant
Glycine
Arginine
Glutamate
Glycine
GENETIC CODE
1.
The genetic code consists of triplets of nucleotides called
amino acid.
TTT Phe
TCT Ser
TAT Tyr
TTC Phe
TCC Ser
TAC Tyr
TTA Leu
TCA Ser
TAA Och
TTG Leu
TCG Ser
TAG Amb
CODONS.
Each codon encodes one
TGT
TGC
TGA
TGG
Cys
Cys
Umb
Trp
CTT
CTC
CTA
CTG
Leu
Leu
Leu
Leu
CCT
CCC
CCA
CCG
Pro
Pro
Pro
Pro
CAT
CAC
CAA
CAG
His
His
Gln
Gln
CGT
CGC
CGA
CGG
Arg
Arg
Arg
Arg
ATT
ATC
ATA
ATG
Ile
Ile
Ile
Met
ACT
ACC
ACA
ACG
Thr
Thr
Thr
Thr
AAT
AAC
AAA
AAG
Asn
Asn
Lys
Lys
AGT
AGC
AGA
AGG
Ser
Ser
Arg
Arg
GTT
GTC
GTA
GTG
Val
Val
Val
Val
GCT
GCC
GCA
GCG
Ala
Ala
Ala
Ala
GAT
GAC
GAA
GAG
Asp
Asp
Glu
Glu
GGT
GGC
GGA
GGG
Gly
Gly
Gly
Gly
2.
Because there are 64 codons and only 20 amino acids to be encoded, the code is REDUNDANT.
Up to six codons may encode the same amino acid and often the nature of the third nucleotide
in the codon is either unimportant or only important in so far as it is a purine or a pyrimidine.
3.
The single methionine codon (AUG) is also a START SIGNAL and sets the READING FRAME.
4.
The AMBER (UAG), OCHRE (UAA), and UMBER (UGA) codons encode no amino acid and are
therefore called NONSENSE CODONS. They act as STOP SIGNALS.
5.
The code is (almost) universal.
6.
The non-overlapping, non-punctuated, triplet nature of the code was proved by CRICK &
BRENNER with genetic crosses of FRAME-SHIFT MUTANTS.
7.
The code was finally cracked by NIRENBERG, KHORANA and other workers with artificial RNA.
tRNA AND TRANSLATION
1.
Ribosomes consist of three RNA strands bound to protein subunits and bring about protein
synthesis by joining amino acids together.
2.
Amino acids are brought to the ribosome for joining in the correct order by charged transfer
RNA (tRNA) molecules which find the appropriate codons and sit in two sites of the ribosome
3.
Particular amino acids are always joined to the 3' end of a particular tRNA molecule by
AMINO-ACYL-tRNA-SYNTHETASE.
4.
tRNA is tightly folded to produce three loops and the amino-acyl arm.
5.
The middle loop (the anticodon loop) contains three nucleotides complementary to a codon for
the amino acid with which the tRNA is charged. These form the ANTICODON.
6.
The first nucleotide of the anticodon (the WOBBLE POSITION) pairs only semi-specifically. Some
anticodons, therefore, pair with more than one codon.
BACTERIAL RNA TRANSCRIPTION
1.
RNA is copied from DNA by RNA POLYMERASE.
2.
RNA polymerase uses RIBONUCLEOTIDE TRIPHOSPHATES as precursors.
3.
RNA polymerase consists of five main protein subunits. A fifth subunit (the  subunit) is
involved in initiation of a new RNA strand.
4.
RNA polymerase does not need a primer to begin synthesis but needs a specific sequence in
the DNA called a PROMOTER.
5.
A promoter has two short conserved sequences 10bp and 35bp to the 5' side of the
transcription start point. Mutations in these destroy promoter activity.
6.
RNA synthesis stops at a
DNA.
TERMINATOR,
where the RNA is caused to fold up and drop off the
THE lac OPERON
1.
FRANÇOIS JACOB & JACQUES MONOD proposed the OPERON HYPOTHESIS. This says that several
cistrons are clustered into OPERON.
2.
The whole operon is transcribed as one mRNA.
3.
The lac operon is controlled by a REPRESSOR protein produced by the lacI gene.
4.
In the absence of lactose, the repressor binds to the OPERATOR (lacO) cistron, which lies
between the promoter and the other cistrons and physically blocks the way of RNA
polymerase.
5.
Lactose binds to the repressor and inactivates its operator binding activity. RNA polymerase
can now bind to the promoter (lacP) and transcribe through the operator and other cistrons.
P
I
P O
Z
Y
A
DNA
RNA
polymerase
Transcription
mRNA
(polycistronic message)
Active repressor
binds to operator
protein
LacI
repressor
lactose
-galactosidase
enzyme
lactose
permease
Inactive repressoreffector complex
lactose
transacetylase
.
lacI mutants are expressed all of the time (CONSTITUTIVELY) because they have no active
repressor. They are recessive.
7.
lacOc mutants are also constitutive but they are dominant. They are called OPERATOR
CONSTITUTIVE MUTANTS because the operator has mutated so that the repressor can no longer
recognise it.
8.
Many cistrons are organised into operons - not just the lac operon.
Cross feeding in Escherichia coli (E. coli) tryptophan-requiring mutants
Several types called TrpA, TrpB, TrpC, TrpD, TrpE
Seed these on an agar plate containing no Tryptophan
Area seeded
with bacteria
(broken line)
TrpA
TrpB
TrpC
Agar
Bacterial
growth
Test nutritional requirements
TrpB mutant needs tryptophan (nothing else will do)
TrpA mutant needs tryptophan or indole
TrpC mutant needs tryptophan or indole or indole glycerol phosphate (IGP)
Biosyntrhetic pathway
   CDRP

IGP

indole

Other Trp
enzymes
TrpC
enzyme
TrpA
enzyme
TrpB
enzyme
Other trp
genes
trpC
gene
trpA
gene
trpB
gene
tryptophan