Chapter 8.8

Question 1

Explain the Sanger method of DNA sequencing

Answer 1

Also called SANGER’S CHAIN TERMINATOR TECNIQUE”

It uses the DNA polymerase enzyme to generate a series of DNA fragments that are complimentary to the DNA being sequenced.

4 solutions contain:

1. A primer
2. An unknown DNA sequence
3. Replication Enzymes
4. Normal nucleotides (A, C, T, G)
5. And a small amount of a terminator nucleotide ( A or C or T or G)
6. To sequence the DNA, it must first be separated into two strands.
7. The strand to be sequenced is copied using chemically altered bases.
8. These altered bases ( terminator) cause the copying process to stop each time one particular letter is incorporated into the growing DNA chain.
9. This process is carried out for all four bases,
10. The result is a group of fragments that differ in length from one another by one end base.
11. Once a collection of such pieces is generated a technique called ELECTROPHORESIS can be used to separate the fragments by size
12. Reading the end bases in order by size reveals the sequence of the compliment the deriving the the original /unknown sequence is easy.

5 the fragments are put together like a jigsaw to reveal the sequence of the original piece of DNA.

Question 2

DNA MICROARRAYS

Answer 2

( also called DNA chip)

. it offers a way to sequence DNA on smaller scale

Question 3

POLYMERASE CHAIN REACTION (PCR)

Answer 3

It replicates millions of copies of a DNA sequence of interest ( in a test tube)

Question 4

The requirements of PCR replication

Answer 4

1. Target DNA sequence to be replicated
2. Taq polymerase ( a heat stable DNA polymerase derived from bacterium that occupies hot springs)
3. 2 types of laboratory made DNA primers that are complimentary to the sequences known o occur at each end of the target sequence. The PRIMERS are necessary because DNA polymerase can only attach to an existing strand.
4. a supply of 4 types of DNA nucleotides

Question 5

Why are Primers necessary

Answer 5

he PRIMERS are necessary because DNA polymerase can only attach to an existing strand.

Question 6

The steps of PCR

Answer 6

1. heat ( from a thermal cycler) separates the 2 strands of the target Dna
2. The temperature is then lowered - short primers attach to the separated target strands by complementary pairings.
3. Tag DNA POLYMERASE adds nucleotides to the primers and builds sequences complimentary to the target
   sequence
4. the heat is raised again: The newly synthesized strands act as templates in the next round of replications which are are separated.
5. The number of pieces of DNA doubles with every round of PCR

Question 7

What is PCR’s greatest strength

Answer 7

it works on tiny samples: a trace amount of dan from a few skin cells ar a strand of hair can filed enough material for profiling.

Question 8

How is PCR used in forensics

Answer 8

to amplify DNA needed to establish genetic relationships, identify remains, convict criminals, and exonerate the falsely accused

Question 9

What is PCR greatest sensitivities

Answer 9

Its it’s extreme sensitivity. Contimination from stray DNA can lead to a false result

Question 10

DNA PROFILING

Answer 10

uses just the most variable parts of the genome to detect genetic differences between individuals

Question 11

SHORT TANDEMS REPEATS

Answer 11

one approach to DNA profiling that, which are sequences of few nucleotides that are repeated in noncoding regions of Dna( silent DNA)

Question 12

The uniqueness of short tandem repeats

Answer 12

For each of the accepted 13str sites each person will have different/unique number of repeats for each nucleotide

Question 13

How is a DNA profile generated

Answer 13

1.DNA is extracted from a person’s cell
2.PCR is used to amplify the DNA at each of the 13 STR sites.
Electrophoresis is then used to determine the number of repeats at each site
by measuring the length difference in the fragments

Question 14

what are the statistics that of the chance of any two unrelated individuals have the same pattern in all of the 13 STR markers

Answer 14

one in 250 trillion

Question 15

From who is mitochondrial DNA inherited . How is it useful in DNA profiling

Answer 15

only from one’s mother. Although it is not useful to distinguish amon siblings, however it is useful to verifying the relationship between woman and child

Question 16

How is father son relationship verified

Answer 16

STRS on the Y chromosome

Question 17

. How do researchers use the Sanger method and DNA microarrays to deduce a DNA sequence?

Answer 17

The Sanger method uses 4 test tubes with the materials needed to replicate many copies of DNA. Included in each test tube are special terminator nucleotides which result in replicated strands of various lengths indicating the positions of A, C, T, and G in the unknown DNA. When the replicated strands are added to a gel electrophoresis plate the complimentary DNA sequence can be determined. Microarrays also determine unknown DNA sequences as the small segments bind to complimentary sequences on the microarray chip. A computer then pieces together the overlaps in sequences to determine the original sequence.

Question 18

How do target DNA, primers, nucleotides, and Taq DNA polymerase interact in PCR

Answer 18

The primers attach to both ends of the target DNA to give the DNA polymerase the necessary nucleotides. The Taq form of the polymerase is not denatured by the heating process that occurs in PCR, and the nucleotides provide the raw material the polymerase needs to replicate the strand.

Question 19

Why is PCR useful

Answer 19

PCR creates many copies of a tiny DNA sequence for analysis, especially in forensics. It can also be used to increase the amount of DNA recovered from a microorganism to aid in diagnosis or from a fossil to aid in evolutionary classification

Question 20

What are STRs, and how are they used in DNA profiling

Answer 20

Short tandem repeats are a series of just a few repeating nucleotides that occur in noncoding portions of DNA. Each individual varies in the number of repeats they possess, so by combining analysis of several STR sites between individuals, a DNA profile can be produced.

Question 21

Why does mitochondrial DNA provide different information from nuclear DNA?

Answer 21

Mitochondrial DNA is much shorter than nuclear DNA and exists in many copies between all the mitochondria in a cell. It is inherited only from the female parent and does not contain a recombination of male and female nuclear DNA. For this reason it cannot be used to distinguish between siblings.