video #2 dna: the blueprint of life name the technology used in the movie jurassic park. where did...
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Video #2 DNA: The Blueprint of LifeName the technology used in the movie Jurassic Park.Where did Meissner extract the “nuclein” material that later was identified as
DNA?How did Hershey & Chase separate the virus from its bacterial host? How did they
trace (track) the DNA and protein?What did x-ray crystallography reveal about DNA?What purpose do enzymes serve in the replication process?Segment #2:Name the disorder that Andrew and his sister inherited. What were the major
symptoms of this disorder?How can this genetic defect be treated? Name the gene that is defective.How can a gene be transported and carried to a cell?What is a vector? Give an example.What purpose do restriction enzymes serve? What about ligase?What does PCR stand for?Segment #3:What is the first step of gene therapy? How long would all of the DNA contained in all of the chromosomes in a human
cell be if they were connected end to end? Which chromosome consists of 5% of all the genes in the human genome?
Protein Synthesis: Chapter 17Bridging the gap between Genotype & Phenotypes
(proteins are thought to be that link)Trace the Flow of Information from Gene to Protein
Key Topics:• Garrod• Beadle & Tatum• Transcription (nucleus)• Processing mRNA• Translation (cytoplasm)• Completed polypeptide (protein)
Introductory Questions #31) Name the substance that accumulates in a person’s urine causing
alkaptonuria. 2) Why did Beadle and Tatum use breadmold spores to determine
that one gene forms one polypeptide allowing for the first metabolic pathway to be defined?
3) Transcribe & Translate the following sequence of DNA by determining the nucleotide sequence for mRNA, the anticodon for tRNA, and the overall amino acid sequence:
TACTCAGGACCTGCAACGATTmRNA: ???????????????????????????????Amino acids Sequence: ???????????????????????????????Anticodon: ???????????????????????????????
4) How does the DNA and amino acid sequences differ from a person with sickle cell anemia and a person with normal hemoglobin in their RBC’s? (pg. 328)
5) When mRNA is “processed” what is taken out (spliced)?
Key Discoveries• Miescher (isolated “nuclein” from soiled bandages) 1869• Garrod (Proteins & inborn errors) 1902• Sutton (Chromosome structure) 1903• Morgan (Gene mapping) 1913• Sumner (Purified Urease, showed it to be an enzyme) 1926• Griffith’s Experiment (Transforming Principle) 1928• Avery, McCarty, and Macleod 1944• Chargaff (Base pairing & species specific) 1947• Hershey and Chase 1952• Pauling, Wilkins, and Franklin 1950’s• Watson and Crick 1953• Meselson & Stahl 1956
Archibald Garrod (1902-1908)• First to suggest that genes dictate phenotypes through
enzymes and their metabolic, catalytic properties.• Studied a rare genetic disorder: Alkaptonuria• Thought to be a recessive disorder• Tyrosine is not broken down properly into carbon dioxide
and water.• An Intermediate substance: “Homogentisic acid”
accumulates in the urine turning it BLACK when exposed to air.
• An enzyme was thought to be lacking• A genetic mutation was thought to be the cause “An Inborn Error of Metabolism”
Metabolic Pathway for the breakdown of Tyrosine
Tyrosine↓
Hydroxyphenylpyruvate↓
Homogentisic acid
Alkaptonuria Maleyacetoacetate Inactive (lacking) enzyme (active ↓ enzyme)
CO2 & H2O
Garrod’s Conclusion
• A mutation in a specific gene is associated with the absence of a specific enzyme.
• Led to the idea of:“One gene, One Enzyme”
• Not validated until Beadle & Tatum’s work in the 1940’s with Neurospora (breadmold)
James Sumner (1926)
• Isolated the enzyme “Urease”
• First to identify an enzyme as a protein
• First to crystallize an enzyme
• Awarded the Nobel prize in 1946 in chemistry for his crystallization of an enzyme.
• Studies of inherited metabolic disorders first suggested that phenotype is expressed through proteins
• Studies of the bread mold Neurospora crassa led to the one gene-one polypeptide hypothesis (Beadle & Tatum)
Figure 10.6B
George Beadle & Edward Tatum
• Discovered the “One Gene, One Enzyme” Principle• Analyzed mutations that interfered with a known metabolic
pathway• Organism they chose to work with: Neurospora
(breadmold)
-Grows easily
-Grows as a haploid: (no homologs)
-Mutants are easily identified: Dominant allele won’t be expressed
• Neurospora (wild type) can grow easily in only: salt, sugar, & Biotin (vitamin) = “Minimal Medium”
George Beadle & EdwardTatum cont’d
• Breadmold spores were bombarded with x-rays & UV• Separated out the survivors: Mutants• Haploid spores were crossed, grown in a variety of media to
determine what kind of mutation was occurring• Mutants (had diff. nutritional needs vs. the wild types) –were
unable to make certain organic molecules: amino acids, lipids, etc. when grown on the minimal media.
• Mutants cold grow on complete growth medium• These mutants were grown in minimal media with one added
nutrient to determine the metabolic defect.• Ex. Arginine was one nutrient that supported growth of the
mutants. • Conclusion: These mutants had defective biochemical pathways
that allowed for the synthesis of arginine.
• **They examined the effect of the mutation instead of identifying the enzyme.
Beadle & Tatum’s Conclusion
“One Gene affects One Enzyme”Later Revised
“One Gene affects One Protein”
Later Revised
“One Gene affects One Polypeptide Chain”
• The information constituting an organism’s genotype is carried in its sequence of bases
THE FLOW OF GENETIC
INFORMATION DNA → RNA → PROTEIN
• The “words” of the DNA “language” are triplets of bases called codons– The codons in a gene specify the amino acid
sequence of a polypeptide
Genetic Information Written in Codons is Translated into Amino acid Sequences
• Virtually all organisms share the same genetic code
• 1st codon determined was “UUU” by Marshal Nirenberg in 1961.
• All of the codons were determined by the mid 1960’s
The Genetic Code Dictionary
Figure 10.8A
Figure 10.7
DNA molecule
Gene 1
Gene 2
Gene 3
DNA strand
TRANSCRIPTION
RNA
Polypeptide
TRANSLATIONCodon
Amino acid
Introductory Questions #31) Name the substance that accumulates in a person’s urine causing
alkaptonuria. 2) Why did Beadle and Tatum use bread mold spores to determine
that one gene forms one polypeptide allowing for the first metabolic pathway to be defined?
3) Transcribe & Translate the following sequence of DNA by determining the nucleotide sequence for mRNA, the anticodon for tRNA, and the overall amino acid sequence:
TACTCAGGACCTGCAACGATTmRNA: ???????????????????????????????Amino acids Sequence: ???????????????????????????????Anticodon: ???????????????????????????????
4) How does the DNA and amino acid sequences differ from a person with sickle cell anemia and a person with normal hemoglobin in their RBC’s? (pg. 328)
5) When mRNA is “processed” what is taken out (spliced)?
Figure 10.7
DNA molecule
Gene 1
Gene 2
Gene 3
DNA strand
TRANSCRIPTION
RNA
Polypeptide
TRANSLATIONCodon
Amino acid
Protein Synthesis: overview
• One gene-one enzyme hypothesis (Beadle and Tatum)
• One gene-one polypeptide (protein) hypothesis
• Transcription: synthesis of RNA under the direction of DNA (mRNA)
• Translation: actual synthesis of a polypeptide under the direction of mRNA
4) How many sites are present in the ribosome? Name the enzyme that is used to attach an amino acid to the tRNA molecule.
• An exercise in translating the genetic code
Figure 10.8B
Startcodon
RNA
Transcribed strand
StopcodonTranslation
Transcription
DNA
Polypeptide
• In transcription, the DNA helix unzips
– RNA nucleotides line up along one strand of the DNA following the base-pairing rules
– The single-stranded messenger RNA peels away and the DNA strands rejoin
RNA polymerase
DNA of gene
PromoterDNA Terminator
DNAInitiation
Elongation
Termination
Area shownin Figure 10.9A
GrowingRNA
RNApolymerase
Completed RNA
Figure 10.9B
Transcription
• Occurs in the nucleus
• RNA Polymerase II is needed
-Adds nucleotides to the 3’ end only
-Eukaryotes have three types vs. Bacteria
with only one type
• Elongation occurs from 5’ 3’ direction
• TATA Box : initiation site for the attachment of RNA polymerase
• 3 Steps: Initiation Elongation Termination
Transcription: Initiation
• RNA Polymerase II binds to the “Promoter” region on the DNA RNA Polymerase recognizes this region because of the “TATA” box
• TATA Box is (upstream about 25 nucleotides from starting pt)
• Other proteins also are needed: – Transcription factors
• These proteins must bind to the DNA first before RNA Polymerase can bind and begin Transcription.
• No Primer is needed, mRNA can made from scratch
Transcription produces genetic messages in the form of RNA
Figure 10.9A
RNApolymerase
RNA nucleotide
Direction oftranscription
Newly made RNA
Templatestrand of DNA
Transcription: Elongation
• DNA is untwisted (hydrogen bonds are broken)
• About 10 base pairs are exposed
• Nucleotides are are added to the 3’ end of the growing mRNA molecule
• Proceeds at a rate of: 60 nucleotides/sec
Transcription: Termination
• Termination site is reached by RNA Polyermase
• In Eukaryotes “AATAAA” is the signal
• In Bacteria Translation can occur as it is released from the first transcription event
• Final mRNA molecule is made consisting of “Coded” and “Non-coded” regions
• Noncoding segments called introns are spliced out
• A cap and a tail are added to the ends
• http://highered.mcgraw-hill.com/sites/0072437316/student_view0/chapter15/animations.html#
Eukaryotic RNA is processed before leaving the nucleus
Figure 10.10
DNA
RNAtranscriptwith capand tail
mRNA
Exon Intron IntronExon Exon
TranscriptionAddition of cap and tail
Introns removed
Exons spliced together
Coding sequence
NUCLEUS
CYTOPLASM
Tail
Cap
mRNA Structure• 1) 5’ cap: modified guanine; protection; recognition site
for ribosomes• 2) 3’ tail: poly(A) tail (adenine); protection;
recognition; transport• 3) RNA splicing: involves introns & Exons• Exons (expressed sequences) retained• Introns (intervening sequences)
-These are spliced out / spliceosome
Animated View of Transcription
• http://highered.mcgraw-hill.com/sites/0072437316/student_view0/chapter15/animations.html#
Translation
• Occurs in the Cytoplasm
• Key molecules and structures include:– mRNA– tRNA– Ribosome (30s and 40s subunits)– Free floating amino acids– Endoplasmic reticulum
• In the cytoplasm, a ribosome attaches to the mRNA and translates its message into a polypeptide
• The process is aided by transfer RNAs
Transfer RNA molecules serve as interpreters during translation
Figure 10.11A
Hydrogen bond
Amino acid attachment site
RNA polynucleotide chain
Anticodon
Translation: Transfer RNA (tRNA)-Pg. 273
mRNA from nucleus is ‘read’ along its codons by tRNA’s anticodons at the ribosome
tRNA – has the anticodon and amino acid attached
• Each tRNA molecule has a triplet anticodon on one end and an amino acid attachment site on the other
Figure 10.11B, C
Anticodon
Amino acidattachment site
Translation- the Ribosome
rRNA site of mRNA codon & tRNA anticodon coupling
P site holds the tRNA carrying
the growing polypeptide chain
A site holds the tRNA carrying
the next amino acid to be added to the chain
E site discharged tRNA’s
Animated View of Transcription
• http://highered.mcgraw-hill.com/sites/0072437316/student_view0/chapter15/animations.html#
Video #4:Proteins-Building Blocks of Life
• Name the structures identified by Dr. James Lake that has helped him to trace the hereditary path of life back to the first cell.
• What type of therapy is suggested by Dr. Richard Firtel that may provide long term help for patients suffering from sickle cell anemia?
• In the third segment what type of organism is profiled? How do prokaryotic cells switch protein production on and off?
Be sure to write the title for all three segments and list five key statements for each segment.
Translation
• Initiation~ union of mRNA, tRNA, small ribosomal subunit; followed by large subunit
• Elongation~ •codon recognition •peptide bond formaton •translocation
• Termination~ ‘stop’ codon reaches ‘A’ site
• Polyribosomes: translation of mRNA by many ribosomes (many copies of a polypeptide very quickly)
• Mutations are changes in the DNA base sequence– These are caused by errors in DNA replication or by
mutagens– The change of a single DNA nucleotide causes
sickle-cell disease
Mutations can change the meaning of genes
DNA Mutations & Modifications• Point mutations
-changes in 1 or a few base pairs in a single gene
-most common
• Base-pair substitutions: •silent mutations no effect on protein
•missense ∆ to a different amino acid (different protein)
•nonsense ∆ to a stop codon and a nonfunctional protein
• Base-pair insertions or deletions:
-additions or losses of nucleotide pairs in a gene; alters the ‘reading frame’ of triplets
-frameshift mutation
• Mutagens: physical and chemical agents that change DNA
• Types of mutations
Figure 10.16B
mRNA
NORMAL GENE
BASE SUBSTITUTION
BASE DELETION
Protein Met Lys Phe Gly Ala
Met Lys Phe Ser Ala
Met Lys Leu Ala His
Missing
Figure 10.16A
Normal hemoglobin DNA
mRNA
Normal hemoglobin
Glu
Mutant hemoglobin DNA
mRNA
Sickle-cell hemoglobin
Val
Key Topics and Text Pgs to ReviewTopic Pgs.
Bacteria: Genetic recombination 346-350Plasmids & ConjugationTransformation (Lab #8)
Transposons: 351-352Lac Operon System 353-356Regulating Gene ExpressionViruses: DNA, RNA (retroviruses) 338-342Lytic & Lysogenic Cycle 337-339
Introductory Questions #41) Name the two scientists that discovered the Lac
operon system.2) How are repressible operons different from inducible
operons? Give an example of each.3) What is the difference between an operator and a
promoter?4) Why are transposons called “jumping genes”? What
purpose do the insertion sequences play?5) Name three example of a virus that has DNA as its
genetic material and three examples of Viruses with RNA as its genetic material.
6) Briefly explain what a vaccine is and what it does.
Repressible Operons (trp operon)• Usually “ON” - to turn OFF:
– Co-repressor needs to bind to an inactive repressor and activate it
– RNA Polymerase then cannot bind and transcribe mRNA
Ex. trp operon is a repressible operon: -trancription is usually on-inhibited only by tryptophan
(corepressor)
Trp Operon when Tryptophan is Absent
http://highered.mcgraw-hill.com/sites/0072437316/student_view0/chapter18/animations.html#
INDUCIBLE Operons (ex. lac operon)• Usually “OFF” - to turn ON:
– INDUCER needs to bind to an active repressor and inactivate it
– RNA Polymerase can then bind and transcribe mRNA
Ex. Lac operon is an inducible operon
Lac Operon• Lactose ONLY used when glucose is not present
in large quantities• When glucose is present, cAMP levels are low,
cAMP cannot bind to CAP and initiate enzyme production
Inactive Repressor-Lactose Presenthttp://highered.mcgraw-hill.com/sites/0072437316/student_view0/chapter18/animations.html
Lac Operon• In absence of glucose, cAMP levels are
HIGH, binding to CAP can occur
• Beta-Galactosidase is made