Unit 3 - Biotechnolgy pt 1 (manipulating and cloning DNA)

Question 1

Genetic Engineering

Answer 1

the intentional production of new genes and alteration of genomes by the substitution or introduction of new genetic materil
ie. human insulin

Question 2

Genetic engineering example

Answer 2

• human gene that code for insulin is inserted into EE.coli plasmid
• E.coli transcribes and translates the piece of human DNA to make the human protein insulin which is then harvested from the bacteria (safflower)

Question 3

Recombinant DNA

Answer 3

a DNA strand created using DNA pieces from two or more sources

Question 4

Restriction Enzymes

Answer 4

act like molecular scissors and cut DNA at specific locations and recognizes a specific sequence of nucleotides on DNA strand

Question 5

recognition site

Answer 5

these r the specific location that r 4 to 8 pairs long and are palindromic (both strands)

Question 6

Restriction fragment

Answer 6

are the fragment produced when a DNA strand is cut by a restriction enzyme

Question 7

Restriction enzyme

Answer 7

each enzyme is cut at only one recognition site and in only one direction

Question 8

types of ends

Answer 8

blunt and sticky

Question 9

blunt

Answer 9

is cut made straight across the strand

Question 10

sticky

Answer 10

cut in a zigzag. scientist prefer to work with sticky ends since they are easier to rejoin w other fragments cut with the same enzyme

Question 11

History of Restriction enzymes

Answer 11

Dr. Hamiliton smith of johns hopkins discovered them in 1970s
since then 2500 have be catalogued with about 200 diff target sequences

Question 12

Restriction enzymes naming process

Answer 12

named after its cell origin plus a roman numeral if more than one restriction enzyme has been isolated from species

Question 13

DNA ligase

Answer 13

is the enzyme that is used to join cut strans of DNA

Question 14

T4 DNA ligase

Answer 14

works well with blunt ends
-less efficient more likely to fail

Question 15

Methylases

Answer 15

enzymes which add a methyl group to adenine to cytosine base within the recognition sequence preventing enzyme from hydrolyzing cells own DNA

Question 16

competent cell

Answer 16

is a cell that is able to take up foreign DNA from surroudings

Question 17

Host cell

Answer 17

a cell that has taken up a foreign plasmid or virus and has used its cellular machinery to express foreign DNA

Question 18

Vector

Answer 18

DNA mol used as a vehicle to transfer foreign genetic material into cell
ie. plasmid

Question 19

cloned gene

Answer 19

and identical copy of an original target gene that can be made by introducing the target gene into a host cell and having a copie

Question 20

Plasmids

Answer 20

common vector used for transferring foreign genetic material into a cell

Question 21

Restriction map

Answer 21

a diagram that shows the relative location of all the know restriction enzyme recognition sites on a particular plasmin and the distance, in to between the sites ( helps pick the correct plasmid to use a host for recombinant DNA procedure)

Question 22

Transformation

Answer 22

under the specific conditions, plasmid enter bacterial cells multiply and express the foreign gene that has been inserted into the plasmid genome

Question 23

DNA cloning

Answer 23

occurs after the use of restriction enzymes and ligation and transfers newly made plasmids to bacteria. after tranformation, bacteria r selected on antibiotic plates. bacteria with a plasmid r antibiotic forming a colony

Question 24

colonies

Answer 24

with the right plasmid can be grown to make large cultures of identical bacteria which r used to produce plasmin or make protein

Question 25

Transformation; some bacteria don't take up plasmids naturally...

Answer 25

- instead these bacteria r placed in solution w calcium chloride and recombidiant DNA plasmids

Question 26

heat shock

Answer 26

through this process of heating and cooling the bacteria cell membrane is disrupted and the plasmid enters

Question 27

steps of bacterial transformation (1)

Answer 27

specially prepared bacteria r mixed w recombinant DNA filled plasmids

Question 28

steps of bacterial transformation (2)

Answer 28

bacteria r given a heat shock which causes some of them to take up plasmid

Question 29

steps of bacterial transformation (3)

Answer 29

plasmids used in cloning contain an antibiotic resistance gene. thus all of the bacteria r placed on an antibiotic plate

Question 30

steps of bacterial transformation (4)

Answer 30

bacteria without a plasmid die. each bacterium with a plasmid gives rise to a cluster of identical plasmid containing a colony

Question 31

steps of bacterial transformation (5)

Answer 31

several colonies check to identify one w the right plasmid (PCR)

Question 32

steps of bacterial transformation (6)

Answer 32

a colony containing the right plasmid grown in bulk and used for plasmid or protein production