The Next Three Weeks: Today: Sanger Sequencing Central Dogma
Overview Mutation Unpacking Central Dogma Transcription (Mar 28)
RNA Processing (Mar 28) Translation (Mar 28 / Apr 2) [+ Sculpting]
Regulation of Gene Expression + Trivia (Apr 4) Tutorial (Apr 5)
Review (Apr 9) EXAM 3 (Apr 11)
Slide 2
Sanger Sequencing
Slide 3
dideoxyNucleoside TriPhosphate
Slide 4
Slide 5
2011 Pearson Education, Inc.
Slide 6
Slide 7
Slide 8
Slide 9
Slide 10
Central Dogma Overview
Slide 11
2011 Pearson Education, Inc.
Slide 12
The Language of Genetics DNA Language vs. Protein Language
Discuss: What does it mean to say that DNA is written in one
language and Proteins are written in a different language?
Slide 13
DNA Language 3 GTGCA 5 5 CACGT 3 The language of DNA is written
in nucleotides.
Slide 14
Protein Language A protein is a chain of amino acids.
Slide 15
Connecting the Languages Transcribing versus Translating
-Transcribe means: to rewrite Transcription Example: 3 parts
gravel, 2 parts sand, 1 part cement.
Slide 16
An Academic Transcript: A copy of your grades
Slide 17
Connecting the Languages Transcribing versus Translating
-Transcribe means: to rewrite -Translating means: to change
language Translation Example: 3 parts gravel, 2 parts sand, 1 part
cement. 3 partes de grava, 2 partes de arena, 1 parte de
cemento.
Slide 18
The Language of Genetics DNA Language vs. Protein Language DNA
is written in the language of nucleotides. A message on DNA can be
transcribed (copied) onto a piece of mRNA. A message on mRNA can be
translated into a chain of amino acids. Proteins are written in the
language of amino acids.
Slide 19
DNA mRNA Amino Acids Transcribing: Replace Thymine with Uracil.
Transcribe from the template strand.
Slide 20
DNA mRNA Amino Acids 5 CCACTGATAGACCTT 3 3 GGTGACTATCTGGAA 5
mRNA is made using the template strand Template Strand non
-Template Strand
Slide 21
DNA mRNA Amino Acids 5 CCACTGATAGACCTT 3 3 GGTGACTATCTGGAA 5 5
CCACUGAUAGACCUU 3 The mRNA sequence is nearly identical to the
non-template strand. Transcribing complete. Template Strand non
-Template Strand mRNA
Slide 22
2011 Pearson Education, Inc. The Translation Key:
Slide 23
DNA mRNA Amino Acids Non-Templ: 5 CCACTGATAGACCTT 3 Template: 3
GGTGACTATCTGGAA 5 mRNA: 5 CCACUGAUAGACCUU 3 Pro - Leu - Ile - Asp -
Leu
Slide 24
Transcribe/Translate the Following Sequences: 5 CTGCGTGACTGCAAA
3 3 GACGCACTGACGTTT 5 3 CCGACTCACTGATGC 5 3 ATGTTTGAACTACAG 5 5
TACAAACTTGATGTC 3
Slide 25
Important Properties of the Code It is redundant : All amino
acids except two are encoded by more than one codon. It is
unambiguous: One codon never codes for more than one amino acid. It
is nearly universal: With a few minor exceptions, all codons
specify the same amino acids in all organisms. It is conservative:
The first two bases are usually identical when multiple codons
specify the same amino acid.
Slide 26
Mutations changes to the code
Slide 27
Whats a Mutation? A mutation is any permanent change in an
organisms DNA. Mutations result in new alleles!
Slide 28
Point Mutations Point mutations occur when the DNA polymerase
inserts the wrong base into the newly synthesized strand of
DNA.
Slide 29
Point Mutations Point mutations may be as a result of a
substitution or an insertion/deletion and may be: Silent mutations.
Does not change the amino acid sequence of the gene product.
Missense mutations. Result in changes in the amino acid sequence of
the encoded protein. Nonsense mutations. Results in a stop
codon.
Slide 30
Mutations and their impacts
Slide 31
Slide 32
Slide 33
Slide 34
Slide 35
Slide 36
Describing a Mutation: Three Categories: 1.What happened?
Insertion, deletion, substitution. 2.What was the impact on the
protein? Missense, nonsense, silent 3.What was the reading frame?
Frameshift
Slide 37
Two Examples of mutation 1.Beach Mice (from the text) 2.Taste
in peas
Slide 38
Peromyscus polionotus
Slide 39
Fact Sheet: Peromyscus polionotus Common names: Beach Mouse or
Old Field Mouse. Location: Southeastern U.S.A. Habitat: Sand
burrows in dunes or old fields. Home-range: ~1000 m 2 Breeding:
Monogamous pair- bonding. Litters of 2-8 pups, every 30 days.
Lifespan: 9-12 months.
Slide 40
Fur Color:
Slide 41
The mc1r Gene The mc1r gene is located in Chromosome #16 in
mammals. It codes for the MC1R protein that aids in pigment
synthesis.
Single Substitution
5TGCCCACCCAGGGGCCTCAGAAGAGGCTTCTGGGTTCTCTCAACTCCACCTCCACAGCCACCC
CTCACCTTGGACTGGCCACAAACCAGACAGGGCCTTGGTGCCTGCAGGTGTCTGTCCCGGATGG
CCTCTTCCTCAGCCTGGGGCTGGTGAGTCTGGTGGAGAATGTGCTGGTCGTGATAGCCATCACC
AAAAACTGCAACCTGCACTCGCCCATGTATTCCTTCATCTGCTGTCTGGCCCTGTCTGACCTGATG
GTGAGTATAAGCTTGGTGCTGGAGACGGCTATCATCCTGCTGCTGGAGGCAGGGGCCCTGGTGA
CCCGGGCCGCTTTGGTGCAACAGCTGGACAATGTCATTGACGTGCTCATCTGTGGCTCCATGGT
GTCCAGTCTTTGCTTCCTTGGTGTCATTGCCATAGACCGCTACATCTCCATCTTCTATGCATTACGT
TATCACAGCATTGTGACGCTGCCCCGGGCACGACGGGCCATCGTGGGCATCTGGGTGGCCAGCA
TCTTCTTCAGCACCCTCTTTATCACCTACTACAACCACACAGCCGTCCTAATCTGCCTTGTCACTTT
CTTTCTAGCCATGCTGGCCCTCATGGCAATTCTGTATGTCCACATGCTCACCCGAGCATACCAGCA
TGCTCAGGGGATTGCCCAGCTCCAGAAGAGGCAGGGCTCCACCCGCCAAGGCTTCTGCCTTAAG
GGTGCTGCCACCCTTACTATCATTCTGGGAATTTTCTTCCTGTGCTGGGGCCCCTTCTTCCTGCAT
CTCACACTCATCGTCCTCTGCCCTCAGCACCCCACCTGCAGCTGCATCTTTAAGAACTTCAACCT
CTACCTCGTTCTCATCATCTTCAGCTCCATCGTCGACCCCCTCATCTATGCTTTTCGGAGCCAGGA
GCTCCGCATGACACTCAGGGAGGTGCTGCTGTGCTCCTGGTGA 3 -Substituting 1 of
954 nucleotides -Cytosine to a Thymine (Pyrimidine Pyrimidine)
-Changed the mRNA codon from CGC to UGC
Slide 46
2011 Pearson Education, Inc. The Translation Key:
Slide 47
Consequence of Mutation A single nucleotide mutation from a
Cytosine to a Thymine leads to An amino acid change from an
Arginine to a Cysteine Amino Acid Sequence Dark Fur:
MPTQGPQKRLLGSLNSTSTATPHLGLATNQTGPWCLQVSIPDGLFLSLGLVSLVENVL VVIAITKN
R NLHSPMYSFICCLALSDLMVSISLVLETAIILLLEAGALVTRAALVQQLDNVI
DVLICGSMVSSLCFLGVIAIDRYISIFYALRYHSIVTLPRARRAIXGIWVASIFFSTLFITYY
NHTAVLICLVTFFLAMLALMAXLYVHMLTRAYQHAQGIAQLQKRQGSTXQGFCLKGAX
TLTIILGIFFLCWGPFFLHLTLIVLCPQHPTCSCIFKNFNLYLVLIIFSSIVDPLIYAFRSQEL
RMTLREVLLCSW Amino Acid Sequence Light Fur:
MPTQGPQKRLLGSLNSTSTATPHLGLATNQTGPWCLQVSVPDGLFLSLGLVSLVENVL VVIAITKN
C NLHSPMYSFICCLALSDLMVSISLVLETAIILLLEAGALVTRAALVQQLDNVI
DVLICGSMVSSLCFLGVIAIDRYISIFYALRYHSIVTLPRARRAIVGIWVASIFFSTLFITYY
NHTAVLICLVTFFLAMLALMAILYVHMLTRAYQHAQGIAQLQKRQGSTRQGFCLKGAAT
LTIILGIFFLCWGPFFLHLTLIVLCPQHPTCSCIFKNFNLYLVLIIFSSIVDPLIYAFRSQELR
MTLREVLLCSW
Slide 48
Changing 1 amino acid: Arginine: Strongest + charge Very
hydrophilic Cysteine: Not hydrophilic Forms disulfide bonds
Slide 49
Two Examples of mutation 1.Beach Mice -Missense substitution
mutation of one nucleotide C T -Changes one amino acid: Arginine
Cysteine -Changes the function of the MC1R protein 2.Taste in
peas
Slide 50
Mendels Peas
Slide 51
Fact Sheet: Pisum sativum Common names: Field peas. Native to:
Near East Iraq, Turkey, Israel, Palestine, etc. A key crop of the
Neolithic Agricultural Revolution, oldest recorded samples date to
10,000 BCE.
Slide 52
The Evolution of Wrinkled Peas Wrinkled peas taste sweet.
WrinkledRound
Slide 53
Molecular Biology: Central Dogma The R allele of the sbe1 gene
codes for the SBE1 protein. The SBE1 protein is an enzyme that
converts simple starch (amylose) to complex starch
(amylopectin).
Slide 54
Inside the Pea Cell: Sugar and Starch
Slide 55
Slide 56
Slide 57
Slide 58
Sucrose
Slide 59
The Evolution of Wrinkled Peas Wrinkled peas arent just
wrinkled they taste good too! Wrinkled (r allele of sbe1 gene)
Round (R allele of sbe1 gene) G3P (Intermediates) Amylose
Amylopectin G3P (Intermediates) Amylose non-functioning starch
branching enzyme Sucrose functioning starch branching enzyme
Slide 60
The Genetics of Round and Wrinkled Peas
Slide 61
The sbe1 Gene The sbe1 gene has two alleles: Wrinkled Pea Codes
for functional SBE1 protein. Round Pea Codes for non- functional
SBE1 protein.
Slide 62
3549 nucleotides What does the R allele represent? Round
Pea
Slide 63
The R Allele These nucleotides code for a polypeptide protein
of about 920 amino acids in length:
Slide 64
What does the R allele do? How does the SBE protein lead to
round seeds? It gives rise to highly branched starch (amylopectin).
Highly branched starch in the seeds leads to seeds with low water
content. When the seeds dry, they stay round.
Slide 65
The original 3549 nucleotides plus an additional 800
nucleotides What does the r allele represent? Round Pea
Slide 66
Unbranched starch in the seeds leads to seeds with high water
content. They are sweet, but when the seeds dry, they wrinkle. How
does the altered SBE protein lead to round seeds? It gives rise to
unbranched starch only (amylose). What does the r allele do?
Slide 67
The R allele and the r Allele: 800 bp fragment of DNA inserted
R allele genetic code r allele genetic code
Slide 68
Two Examples of mutation 1.Beach Mice -Missense substitution
mutation of one nucleotide C T -Changes one amino acid: Arginine
Cysteine -Changes the structure and function of the MC1R protein
2.Taste in peas -Nonsense insertion mutation of 800 nucleotides.
-Disrupts amino acid production. -Changes the structure and
function of the starch branching enzyme.
Slide 69
Mutations Insertion/Deletion/Substitution Frameshift or no
Frameshift Silent/missense/nonsense