Exam 3 - Lecture 18

Question 1

what is genetic engineering?

Answer 1

deliberate modification of an organism’s genetic information by directly changing the sequence of nucleic acids in its genome

Question 2

what is recombinant DNA?

Answer 2

artificially created DNA sequences resulting from combining 2 strands of DNA together

Question 3

what is cloning?

Answer 3

generating a large number of genetically identical DNA molecules

Question 4

what is biotechnology?

Answer 4

use of biological organisms to form useful products

Question 5

what is industrial microbiology?

Answer 5

use of microbes to manufacture important compounds

Question 6

what are two main reasons you might want to express a foreign gene in a host cell?

Answer 6

• to determine its function
• to purify it

Question 7

what are four recombinant DNA technologies?

Answer 7

• restriction enzymes
• genetic cloning
• PCR
• DNA sequencing

Question 8

what do restriction enzymes (RE) do?

Answer 8

they recognize and bind specific sequences in DNA called recognition sites to cleave DNA

Question 9

where do the following RE cleave DNA?

• Type I
• Type II
• Type III

Answer 9

• Type I: cleave at a defined distance from the recognition site
• Type II: cleave at/around the recognition site
• Type III: cleave at a defined distance from the recognition site

Question 10

cleavage may cause what kinds of ends in the DNA target?

Answer 10

• blunt ends
• sticky ends

Question 11

true or false: many restriction enzyme sites are palindromes.

Answer 11

true

Question 12

what is a vector?

Answer 12

a carrier of foreign DNA

Question 13

what are four types of cloning vectors? which is the most common*?

Answer 13

• phages and viruses
• cosmids
• artificial chromosomes
• plasmids*

Question 14

to clone a gene, __________ _____ is combined with/inserted into a cloning vector.

Answer 14

foreign DNA

Question 15

what are two characteristics of good cloning vectors?

Answer 15

• easy to purify
• replicate autonomously

Question 16

what are three requirements for vectors?

Answer 16

• origin of replication
• selectable marker (antibiotic resistance gene)
• multiple cloning site (MCS)

Question 17

what is an MCS and what is it used for?

Answer 17

• multiple cloning site
• site where cloned gene is inserted into the plasmid vector
• contains many unique RE sites
• may contain selection gene (ex: lacZ)

Question 18

what is the most common bacterial host for insertion of recombinant DNA?

Answer 18

E. coli

Question 19

what is the most common eukaryotic host for insertion of recombinant DNA?

Answer 19

S. cerevisiae

Question 20

what are two mechanisms for artificially induced competence for transformation?

Answer 20

• CaCl2
• electroporation

Question 21

what is PCR?

Answer 21

• polymerase chain reaction
• technique that enables DNA amplification
• rapid synthesis of many copies of a specific DNA fragment
• individual DNA fragments can be specifically amplified from a complex mixture of DNA

Question 22

what determines the specificity of PCR?

Answer 22

• DNA primers (oligonucleotides)

Question 23

what is the PCR cycle?

Answer 23

• DNA is denatured
• primers anneal to target DNA
• target DNA is synthesized (amplified)
• repeat x 34 times

Question 24

PCR causes _____________ apmlification.

Answer 24

exponential

Question 25

what are the uses of PCR?

Answer 25

• simplifies gene coding
• generates DNA fragments for nucleotide sequencing
• may amplify environmental genes without culturing the microbes
• diagnostic purposes

Question 26

what is the most commonly used method for determining DNA sequences?

Answer 26

Sanger DNA sequencing

Question 27

what type of nucleotides does the Sanger DNA sequencing method use to signal termination?

Answer 27

dideoxynucleoside triphosphates (ddNTPs)

Question 28

strand synthesis terminates when a ________ is incorporated in Sanger DNA sequencing.

Answer 28

ddNTP