Biology 2581B Lecture Notes - Lecture 10: Plasmid, Polyacrylamide Gel Electrophoresis, Lac Operon

26 views7 pages

maroonelk470

3 Mar 2013

School

Department

Course

Professor

For unlimited access to Class Notes, a Class+ subscription is required.

Document Summary

Restriction enzymes are double stranded dna endonucleases. Some have blunt ends, some have 3" sticky ends, some have 5" sticky ends: what do they do, types of ends, calculation of average fragment size: 4n. The n is the length of recognition size. Need to analyze dna after it has been cut with restriction enzyme: basic procedure. Run them on agarose or polyacrylamide gel: basis of separation size. Smaller dna fragments run through faster than larger fragments. Dna amplification 1 molecular cloning: how a plasmid vector works. Amplify dna in bacteria or a bacteriophage. Deal with a plasmid vector that has an f factor and a selectable marker: ligation. Add ligase to get them to come together to form chimeric molecules with plasma dna and human dna: transformation, selection. Select for bacteria that have taken up the plasma dna will get a colony of bacteria that have taken up this plasmid. Dna amplification 2: pcr: primers, thermocycling, chain reaction.

Related Questions

1.) A restriction recognition site is repeated 1 times on a circular DNA molecule. If you treat this DNA molecule with this restriction enzyme, how many DNA fragments will you get?

a. 1

b. 2

c. 3

d. Restriction enzyme does not cut a circular molecule

2.) In an agarose gel DNA electrophoresis, the DNA fragments are separated based on their

a. Size

b. Electrical charge

c. Nucleotides sequences

d. Density

e. All of the above

3.) DNA molecules are usually separated using agarose gel rather than polyacrylamide because:

a. Agarose polymerizes faster than acrylamide gel

b. Polyacrylamide gel has pore size larger than the size of DNA molecules

c. Agarose gel do not have pores on tis matrix and can hold large DNA molecules

d. Agarose gel has larger pore size similar to the size of DNA molecules

4.) Restriction endonucleases

a. Randomly recognize and cut DNA sequences

b. Hydrolysis the phosphodiester bond of DNA backbone

c. Recognize specific sequence on double stranded DNA

d. A and B

e. B and C

taupefrog573

View/perform/read ALL THREE of the following prior to answeringthe questions.

http://highered.mcgraw-hill.com/olcweb/cgi/pluginpop.cgi?it=swf::535::535::/sites/dl/free/0072437316/120078/micro10.swf::Stepsin Cloning a Gene (Links to an external site.)

http://www.discoverbiotech.com/wiki/-/wiki/Main/Applications ofCloning (Links to an external site.)

http://www.wiley.com/college/boyer/0470003790/animations/cloning/cloning.htm(Links to an external site.)

Skip to question text.

From the list below, which of the following is the most logicalsequence of steps for splicing foreign DNA into a plasmid andinserting the plasmid into a bacterium?
I. Transformbacteria with recombinant DNA molecule
II. Cutthe plasmid DNA using restriction enzymes
III. Extractplasmid DNA from bacterial cells
IV. Hydrogen-bondthe plasmid DNA to nonplasmid DNA fragments
V. Useligase to seal plasmid DNA to nonplasmid DNA

Flag this Question

Plasmids (or vectors) are important in biotechnology becausethey are

a vehicle for the insertion of recombinant DNA intobacteria.

surfaces for respiratory processes in bacteria.

recognition sites on recombinant DNA strands.

surfaces for protein synthesis in eukaryotic recombinants.

proviruses incorporated into the host DNA

Flag this Question

Plasmids are put into bacterial cells by

restriction enzymes

DNA ligase

binding of cohesive sticky ends

transformation

Flag this Question

Restriction enzymes usually

cut donor DNA evenly so smooth edges result

cut donor DNA but do not affect plasmids

make staggered cuts at specific sequences in DNA in both donorDNA and plasmid

are used in ligating plasmids into bacterial host cells

more than one of the above

Flag this Question

After combining DNA fragments in a cloning experiment, ___ isused to covalently join the DNA segments.

Restriction enzyme

DNA Ligase

Reverse transcriptase

DNA polymerase

Helicase

Flag this Question

It is theoretically possible for a gene from any organism tofunction in any other organism. Why is this possible?

All organisms have ribosomes.

All organisms have the same genetic code.

All organisms are made up of cells.

All organisms have similar nuclei.

All organisms have transfer RNA.

Flag this Question

Skip to question text.

Assume that you are trying to insert a gene into a plasmid andsomeone gives you a DNA sample cut with restriction enzyme X. Thegene you wish to insert from the given sample has sites on bothends for cutting by restriction enzyme Y. You have a plasmid with asingle site for Y, but not for X. Your strategy should be to

cut the plasmid with restriction enzyme X and insert thefragments cut with Y into the plasmid.

cut the plasmid with enzyme X and then insert the gene into theplasmid.

cut the DNA again with restriction enzyme Y and insert thesefragments into the plasmid cut with the same enzyme.

cut the plasmid twice with restriction enzyme Y and ligate thetwo fragments onto the ends of the human DNA fragments cut withrestriction enzyme X.

insert the fragments cut with X directly into the plasmidwithout cutting the plasmid.

Flag this Question

Which of the following is/are false in regard to expressionplasmids (also called expression vectors)?

They are used to make proteins using a cloned gene.

They contain a promotor.

They are the first plasmid type used to clone a gene.

They contain a terminator.

More than one of the above is false.

Flag this Question

What is NOT a potential problem(s) associated with usingbacteria containing a cloned eukaryotic gene (e.g. a human gene) toproduce a functional protein?

If the eukaryotic gene contains introns the bacteria will notremove them and the resulting amino acid sequence will be differentthat that made by a eukaryote.

The bacteria may not fold the protein correctly.

The bacteria may degrade the protein.

All of the above are potential problems.

Flag this Question

Cloning allows for production of proteins in much larger amountsthan occurs in the cells from which the gene is isolated.

True

False

Flag this Question

Question 111 pts

Gene cloning is used to do all of the following except

Make insulin

Making genetically identical animals (e.g. Dolly thesheep)

Make vaccines

Perform Gene Therapy

Making genetically engineered plants

Flag this Question

In your

rubyhedgehog47

1. In a successful PCR reaction, which of the following is true?

a. The template DNA is added in excess of the primers and dNTPs.

b. The annealing temperature must be just below the melting temperature of the primers.

c. The temperature for the denaturation step is 72 C

d. All of the above

2. In our TLC experiment, which of the following had an effect on the migration of the samples?

a. The number of negatively charged phosphate groups

b. The protonation state of ionizable groups on the bases.

c. The size of the molecule.

d. All of the above

3. Which of the following is true of nucleic acid migration in agarose gel electrophoresis?

a.Nicked circular plasmids migrate faster than supercoiled plasmids.

b.Binding of ethidium bromide during electrophoresis speeds up the movement of linear fragments.

c.A higher percentage of agarose will decrease the migration of larger DNA fragments.

d.A lower strength electric field will increase the migration of DNA fragments.

4.What is the purpose of the outgrowth period during transformation?

a.To allow for the cells to take up DNA.

b.To allow for expression of the lacZÎ± and lacZÏ genes.

c.To allow for expression of antibiotic-resistance genes.

d.To prepare cell membranes for electroporation.

5.Which of the following is true of ligase and ligation reactions?

a.Ligase facilitates H-bonds between the sticky ends created by restriction enzyme digestion.

b.Ligase is inactivated at 37 C.

c.DNA fragments must have a 5â PO₄ group for ligase to covalently join them together.

d.Vector must be added in molar excess to insert to favor recombinant plasmid formation.

Biology 2581B Lecture Notes - Lecture 10: Plasmid, Polyacrylamide Gel Electrophoresis, Lac Operon

Document Summary

Get access

Related Documents

Biology 2581B Lecture Notes - Lecture 9: Approximation, Dideoxynucleotide, Chromatography

Biology 2581B Lecture Notes - Lecture 9: Agarose Gel Electrophoresis, Plasmid, Restriction Enzyme

Biology 2581B Lecture Notes - Lecture 9: Agarose Gel Electrophoresis, Nitrocellulose, Selectable Marker

Related Questions