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BIOC 302 Lecture Notes - Lecture 5: Prokaryote, Intron, Consensus Sequence

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greylion206
18 Nov 2015
School
UBC
Department
Biochemistry
Course
BIOC 302
Professor
Michael J.Krisinger

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Document Summary

The next big discussion will revolve around rna metabolism. In this context some major differences between dna and rna were highlighted and so were differences between prokaryotic and eukaryotic cells. The way a gene sequence is reported was explained. Similarities and differences to dna polymerases were briefly explained. The general structure and functionality of a rna polymerase was introduced. To understand the rna pol functionality the basic genetic elements that are required to express a gene have to be understood. The four major such elements were discussed in detail and general examples of typical promoter sequences were given. (check definitions of: promoter, consensus sequence!) The stages and players in the process of initiating transcription were discussed in detail. The difference between closed and open complex and the requirement for promoter clearance were highlighted. The two mechanisms of prokaryotic termination were discussed (rho dependent vs. rho independent termination).

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Related Questions

Chapter 10

1.Outline the history of our knowledge on DNA up to Watson and Crick. What were the main contributions made by each researcher’s key experiment?

2.Explain the setup of the Hershey and Chase experiment, what would the results have been if protein was the genetic material?

3.Draw the structure of a DNA nucleotide, labeling each main component correctly. How does an RNA nucleotide differ?

4.If a section of double stranded DNA contains 19% Adenine, how much Thymine is present?

5.You are a researcher studying the genetic basis of heart attacks and have been working to determine the expression levels of different genes that might contribute to cancer formation. You obtain the DNA methylation status of five genes of interest (the data are shown in the table below). The plus (+) sign indicates the level of DNA methylation; more plus signs correlates with increased methylation levels.Based on this information which genes would you predict to have the highest rate of transcription?

Gene

Methylation levels

1

++

2

+++++

3

+++

4

++

5

+

What are the characteristics of the 3 main DNA forms?

Chapter 11

What are the different types of chromatin?

What are the structures and important roles for telomeres and centromeres?

What are the differences found between eukaryotic chromosomes and mitochondrial?

Chapter 12

Explain each of the different models of replication.

If you grow a culture of bacteria in media with radioactive nucleotides so that all DNA in the cells include radioactive nucleotides and then place the bacteria in new non radioactive media. After two rounds of replication what proportion of the DNA molecules will contain radioactivity?

Summarize the similarities and differences between rolling-circle replication, theta replication and linear eukaryotic replication.

What are the functions of the different DNA polymerases found in eukaryotic cells?

Draw a replication fork and include all key components and orientations. (Leading/lagging strands, DNA helicase, RNA primer and DNA gyrase)

What is the Holliday model of recombination and what are the necessary steps?

Chapter 13

What are the different types of RNA and what roles do they play?

Describe the properties and functions of each of the RNA polymerases and how they differ depending on the organism.

Describe in detail the process and mechanisms of transcription in both prokaryotes and eukaryotes.

Chapter 14

What are the primary purposes of each of the three post transcriptional modifications that occur in eukaryotic cells.

What is alternative splicing and what role does it play in the cell?

How is ribosomal RNA processed after transcription?

How do siRNA and miRNA work, describe/draw out the process in detail.

(tautomerization) The most common form of guanine is the ketoform, not the enol form.

True

False


What term describes the allowed degree of base pair matchingduring annealing, varying from lots of base-pair mismatches allowedto allowing only perfect base-pair matching.

stringency

aneuploidy

polyploidy

quantitative PCR

restriction fragment length polymorphism


I have an individual with a disease we will call SSI-itis,brought on by extreme exposure to cannabis. I do a southern andfind the gene affected. Another of his classmates exhibits theexact same symptoms, but that gene in hiw is wt. A southern in thesecond student finds a mutation in another gene. This is an exampleof

pleitropy

allelic heterogeneity

locus heterogeneity

dominant traits

recessive traits

penetrance


If a person carrying a dominant allele does not demonstrate thephenotype strongly, but still differs from wt, thisdemonstrates?

haploinsufficiency

penetrance

expressivity

monogenic traits

polygenic traits


I find three alleles for a gene in a fruit fly. One allelecreates the ability for the fly to eat graphite. Another allelecreates the ability for the fly to eat coal. The third alleleallows the fly to eat diamond. These alleles are all dominant, andwt cannot eat any of these substances. Which statement below is themost true?

Upon mating a bunch of the flies carrying these alleles, Ishould not find a fly that can eat graphite, coal, and diamond.

I would consider the ability to eat these substances an exampleof pleitropy because they are all carbon.

I would expect the ability to eat diamond to be dominant to theability to eat graphite.

These flies are demonstrating locusheterogeneity.


Expression vectors in prokaryotes do not make functionaleukaryotic gene products in bacteria very well because

the codon sequence for prokaryotes is different than the codonsequence in eukaryotes

there are no disulfide bridges formed in proteins normally madein prokaryotes

prokaryotic expression vectors cannot translate eukaryticsequences

eukaryotic genes have introns, and prokaryotes don't

eukaryotic genes have exons and prokaryotes don't


Which of the following is not used in PCR amplification ofDNA?

Double-stranded DNA template

Oligonucleotide primers

DNA polymerase

DNA ligase

ddNTPs


I create a knockout mouse using the agouti/black fur systemdiscussed in the online lecture and in the book. When I cross theagouti offspring of the originally engineered mouse, I find a ratioof 2 agouti mice to 1 black furred mouse. What is the bestexplanation?

The gene knocked out was recessive.

The gene knocked out was dominant.

The gene knocked out was a lethal gene.

The knockput was integrated into a random spot, and did notknockput the original gene.


A loss of function mutation is usually recessive, but canactually be a dominant mutation because of what effect?

haploinsufficiency

dominant negative mutations

allelic heterogeneity

locus heterogeneity

pleitropy


A genetic carrier of a disease can have a form of the diseasethat only shows intermittent symptoms, while the full homozygousrecessiuve individual would show all of the symptoms of thedisease.

True

False


Wild type, or wt, refers to the genotype that is the typicalgenotype found in nature. Now read that again. Once more. Nowanswer.

Answer

True

False



Which of the following did Erwin Chargaff observe?

A)

Base composition changes as the organism ages.

B)

Base composition of DNA does not vary from one species toanother.

C)

In all species the number of adenines equals the number ofguanines.

D)

In all species A + T = G + C.

E)

DNAs isolated from different tissues of the same species havethe same base composition.

The melting temperature (Tm) of a DNA duplex is:

A)

the temperature at which double-stranded nucleic acids degradeinto free nucleotides.

B)

the temperature at which double-stranded DNA is toosingle-stranded to ever renature back to double strands.

C)

the temperature at which double-stranded DNA is completelymelted.

D)

the temperature at which double-stranded DNA has become 50%single-stranded.

E)

the temperature at which double-stranded DNA begins to melt.

Given your knowledge and the descriptions of each of thesemethods, which of the following does NOT rely on the ability ofnucleic acids to hybridize with each other?

A)

Southern blotting for the detection of specific DNA fragmentsthat have been separated by gel electrophoresis.

B)

Colony blot hybridization to identify a specific sequence from acollection of sequences that have been inserted into bacteria.

C)

Northern blotting for the detection of specific RNA fragmentsthat have been separated by gel electrophoresis.

D)

A DNA oligonucleotide microarray incubated with probes made fromcDNA.

E)

Western blotting for the detection of specific proteins thathave been separated by gel electrophoresis.

Which of the following is a possible reason why DNA uses thymineinstead of uracil?

A)

Cytidine is deaminated to uracil so it would be difficult forany repair mechanism to differentiate between uracil that belongsin the sequence and uracil that resulted from deamination.

B)

Adenine is deaminated to uracil so it would be difficult for anyrepair mechanism to differentiate between uracil that belongs inthe sequence and uracil that resulted from deamination.

C)

When uracil is bound to a deoxyribose it will become morereactive, forming covalent bonds with other bases.

D)

None of the above.

Which of the following is an enzyme used in cloning to breakcovalent bonds?

A)

DNA ligase

B)

Restriction endonuclease

C)

Reverse transcriptase

D)

DNA polymerase I

E)

Alkaline phosphatase

Which of the following could you use to remove the 5' phosphatesafter cleavage of a plasmid with a single type of restrictionenzyme to ensure that the plasmid would not simply fuse backtogether upon addition of ligase?

A)

Reverse transcriptase

B)

Alkaline phosphatase

C)

DNA ligase

D)

DNA polymerase I

E)

Restriction endonuclease

Which of the following would you use to create a cDNA librarythat you would not use for a genomic library?

A)

Alkaline phosphatase

B)

Reverse transcriptase

C)

Restriction endonuclease

D)

DNA ligase

E)

DNA polymerase I

A plasmid vector typically has which of the followingfeatures?

A)

An origin of replication

B)

A selectable marker to ensure that the only bacteria growingcontain the intact or recombinant vector.

C)

Unique restriction sites for insertion of DNA.

D)

Small size to facilitate entry into the cell and biochemicalmanipulation of the DNA.

E)

All of the above are features of plasmid vectors.

Which of the following is FALSE of genomic or cDNAlibraries?

A)

Genes represented in cDNA libraries include control sequencesthat direct transcription.

B)

cDNA libraries contain cDNAs made from mRNA.

C)

Genomic libraries contain fragments of genomic DNA that aregenerated by partial digests with restriction enzymes.

D)

Genomic libraries usually contain large fragments and are likelyto be constructed in YAC or BAC vectors.

E)

cDNA libraries contain sequences from expressed genes only.

Which of the following is NOT a typical component of apolymerase chain reaction (PCR)?

A)

dNTPs

B)

DNA containing the sequences to be amplified

C)

DNA ligase to connect the fragments together

D)

Primers complementary to each end of the sequence to beamplified

E)

Heat stable Taq polymerase

How can a fusion protein be formed?

A)

Site-directed mutagenesis where a fragment is replaced with asynthetic sequence containing the altered DNA sequence.

B)

Two proteins can recombine in the bacterial cell to form a newhybrid protein.

C)

Portions of two different genes are ligated together to create anew hybrid gene. When expressed the gene product is a fusion of thetwo gene products.

D)

Using oligonucleotide-directed mutagenesis to introduce specificmutations that cause the expressed protein to fuse to otherproteins.

E)

A portion of the gene can be removed to form a shorter gene andtherefore a shorter protein.

Why is green fluorescent protein (GFP) so useful in visualizingfusion proteins in eukaryotes?

A)

The GFP protein recognizes and binds to fusion proteins allowingthe location of the fusion protein to be determined.

B)

The reaction is only dependent on the presence of GFP and oneother cofactor.

C)

It is easily cloned and expressed in bacterial cells.

D)

The presence of just a few molecules of GFP fusion proteins canbe sufficient for observing the proteins microscopically allowingthe study of its location and movements in the cell.

E)

It is derived from fire flies, which are easy to cultivate inthe lab.

You wish to determine the cellular location of a protein butunfortunately it is not particularly antigenic (can t develop astrong antibody response). How could you solve this problem?

A)

Create a fusion protein with an epitope tag that can bevisualized by incubation with fluorescently tagged antibody to theepitope.

B)

Create a fusion with green fluorescent protein (GFP).

C)

Express the protein as a fusion with biotin. The addition of GFPwill cause fluorescence.

D)

A and B might work.

E)

A, B, and C might work.

F)

None of the above will work.

A positive result for the yeast two-hybrid analysis means thefollowing:

A)

The proteins that you are studying can be made into fusionproteins, but do NOT interact with each other within the nucleus ofyeast.

B)

A fusion protein containing the Gal4p activation domain, hasbound to RNA polymerase resulting increase transcription ofnumerous genes, including the reporter gene, and formation ofcolonies.

C)

Two fusion proteins, one containing the Gal4p binding sitedomain and one containing the Gal4p activation domain, haveinteracted through their Gal4p domains resulting in transcriptionof the reporter gene.

D)

Two fusion proteins interact with each other and because of theGal4p binding site domain and the Gal4p activation domain containedin each fusion protein, the reporter genes are activated and thecells die resulting in no colonies.

E)

The colonies growing are expressing the two fusion proteins, butthe two proteins have not interacted with each other.

Which of the following is the approximate size of the humangenome?

A)

3 Gm

B)

300 Kb

C)

3 Mb

D)

3 Gb

E)

3 Mm

Which of the following is correct about the structure orlocation of genes?

Question 22 options:

A)

Prokaryotic genes typically possess their own regulatorysequences that only control expression of a single gene.

B)

Eukaryotic genes often encode introns that are spliced outbefore translation.

C)

Eukaryotic genes are mostly located at the telomeres and thecentromeres of chromosomes.

D)

Eukaryotic genes are typically arranged in operons.

E)

Prokaryotic genes often encode exons that are spliced out beforetranslation