BIO 319 Lecture Notes - Lecture 3: Cloning Vector, Plasmid, Real-Time Polymerase Chain Reaction

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coralcicada848

19 May 2017

School

University at Buffalo

Department

Biological Sciences

Course

BIO 319

Professor

Michael Yu, Laura Rusche

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Document Summary

Bio319 genetics - study guide for unit 3, lecture 21 - chapter 20. Recombinant dna technology the use of in vitro molecular techniques to isolate and manipulate different pieces of dna. Recombinant dna molecule molecules that are produced in a test tube by covalently linking dna fragments from two different sources. Gene cloning the technique of isolating and making many copies of a gene using molecular methods, such as pcr or the introduction of a gene into a vector that replicates in a host cell. Vector a small segment of dna that is used as a carrier of another segment of dna. Host cell a cell that is infected with a virus or bacterium or the cell that harbors the vector (plasmid & virus) Expression vector a cloning vector that contains a promoter so that the gene of interest is transcribed into. Rna when the vector is introduced into a host cell.

Related Questions

1- if each E. Coli cell contained fifty copies of the plasmids, how many copies of the plasmid were in the microcentrifuge tube after losing the cells?

2- describe how Jeff could use agarose gel electrophoresis to confirm that the plasmid is recombinant?

3- the stripe rust resistance gene is part of a recombinant DNA molecule consisting of cloning vector linked to wheat DNA. What is a cloning vector? Give two examples of specific DNA molecules routinely used as a cloning vectors?

carminemosquito882

Please answer the following multiple-part question regarding vectored cloning:

a) A cloning vector must _________.

A. be a circular dsDNA molecule

B. contain a replication origin

C. contain an multicloning site (MCS)

D. contain marker genes

b) Vectored cloning is superior to PCR cloning of a gene in terms of __________.

A. time it takes to finish cloning

B. size of DNA that can be cloned

C. amounts of cloned DNA that can be generated

D. ease of isolation of the DNA fragment to be cloned

c) Which of the following is a recombinant DNA molecule?

A. pUC DNA digested by EcoRI

B. Fish insulin gene, PCR amplified and then cut by EcoRI

C. pUC18 DNA and fish insulin DNA ligated by a DNA liagse

D. pUC DNA and fish insulin gene DNA, both cut by EcoRI and mixed

d) You may insert a foreign DNA fragment inside a live cell using each of the following methods except________.

A. inject the DNA inside the cell

B. mix the DNA with the cell and freeze the mixture in liquid nitrogen

C. mix DNA with the cell and momentarily heat the cell then cool down

D. mix DNA with the cell and momentarily zap the cells with a high voltage

E. cover the DNA in a lipid micelle and mix the micelle with the cell

e) Technically, the ampicillin resistance gene is a selectable marker gene but the beta lacZ gene or the green fluorescent protein (GFP) gene is a reporter gene of a cloning vector. What is the difference between a selectable marker and a reporter gene?

A. A selectable marker is a bacterial gene, a reporter gene is a eukaryotic gene

B. Marker genes are used in bacterial system; reporter genes are used in eukaryotic system.

C. The selectable markers help eliminate cells lacking the maker, reporter genes do not offer this option.

1. You have cloned a human gene along with its promoter and ribosome-binding site in a bacterial cloning vector and then introduced the recombinant plasmid into a bacterial host cell that supports the replication and segregation of the cloning vector. Unfortunately, even though you know the whole human gene sequence is present (by DNA hybridization); you cannot get expression of the gene. Which of the following is NOT a reason why there is no expression?

A. The vector cannot replicate

B. The human promoter is not recognized by the bacterial host cell

C. The ribosome-binding site is not recognized by the bacterial host cell

D. The human gene lacks introns, so it is not transcribed by the host bacterium

2. Restriction endonucleases are:

A. sequence-specific

B. able to recognize any 4-, 6-, or 8-base pair sequence

C. always able to produce single stranded tails

D. only able to cleave bacterial DNA

3. Which of the following is NOT a required characteristic of cloning vectors?

A. Genes for replication of vector DNA and segregation to progeny cells

B. Genes for selection of cells carrying the vector (e.g., antibiotic-resistance)

C. DNA sequence for the introduction of foreign DNA where the integrated DNA does not disable the vector

D. None of the above

4. A poly (T) primer is used to produce cDNA from mammalian mRNA. Which of the following would be used to produce cDNA from bacterial mRNA?

A. A poly (T) primer

B. A random DNA sequence primer

C. A poly (A) primer

D. A poly (GC) primer

BIO 319 Lecture Notes - Lecture 3: Cloning Vector, Plasmid, Real-Time Polymerase Chain Reaction

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