DNA sequencing

Question 1

learning objective - outline the steps in automated DNA sequencing via the dideoxy method

Answer 1

-

Question 2

dideoxy sequencing

Answer 2

a method of DNA sequencing that uses dideoxycytidine ribonucleotides to terminate the growth of DNA strands

Question 3

what is the importance of dideoxyribonucleotides?

Answer 3

chemists can synthesize dideoxyribonucleotides (ddNTPS) that are missing the OH group at the 3’ position

Question 4

what do ddNTPS tell us?

Answer 4

Sager reasoned that if ddNTP is added to a growing DNA strand, the strand can no longer go due to lack of the OH group

Question 5

what is the incorporation of ddNTP into a strand called?

Answer 5

chain termination

Question 6

automated DNA sequencing

Answer 6

each type of ddNTP is labeled with a different fluorescent dye

Question 7

How much of the starting DNA do we need for automated DNA sequencing?

Answer 7

DNA to be sequenced is obtained in large amounts, which is done by gene cloning

Question 8

what is the annealing site?

Answer 8

the annealing site is the place in the vector where a primer will bind, which is next to the target DNA that was inserted

Question 9

what is the purpose of automated DNA sequencing?

Answer 9

determine the base sequence of the target DNA next to the annealing site

Question 10

what are the steps of Sanger sequencing?

Answer 10

1) a sample with many copies of the single stranded vector is mixed with primers
2) primer bonds to DNA at annealing site
3) dNTPs are added at high concentrations
4) ddNTPS are added at low concentration
5) DNA polymerase is added

Question 11

sequence ladder

Answer 11

a series of bands on a gel that can be followed in order to determine the base sequence strand of DNA

Question 12

gene mutagenesis

Answer 12

-

Question 13

learning objective

Answer 13

-

Question 14

learning objective

Answer 14

-

Question 15

why is studying mutations important?

Answer 15

geneticists have discovered that many inherited human diseases, like sickle cell disease, involve mutations within specific genes. This provides insight to the function of genes in unaffected individuals

Question 16

site directed mutagenesis

Answer 16

allows a researcher t produce a mutation at a specific site within a cloned DNA segment

Question 17

how is site directed mutagenesis useful?

Answer 17

it can be introduced into a living organism to see how much the mutation affects the expression of a gene, the function of a protein, and the phenotype of the organism

Question 18

why is site directed mutagenesis sometimes called oligonucleotide directed mutagenesis?

Answer 18

an oligonucleotide primer anneals to the template DNA, where the base sequence of this primer is designed. The primer contains a mismatched region that is not complimentary to the DNA, which is where the mutation occurs

Question 19

what components are needed for site directed mutagenesis?

Answer 19

dNTPs, DNA polymerase, DNA ligase

Question 20

what is the final product of this sequencing?

Answer 20

a sequence with a designed mutation that is inserted into bacteria for further investigation

Question 21

crispr-cas technology

Answer 21

a method that uses the components of the crispr-cas system of genome defense found in prokaryotes to introduce mutations into genes

Question 22

-

Answer 22

-

Question 23

-

Answer 23

-

Question 24

-

Answer 24

-

Question 25

-

Answer 25

-

Question 26

-

Answer 26

-

Question 27

21.5 learning objective

Answer 27

-