Unit 5

Question 1

Polymerase chain reaction (PCR)

Answer 1

artificial method of replicating DNA

Question 2

What does PCR involve?

Answer 2

repeated cycling through high and lower temperatures

Question 3

What are the 4 steps of PCR?

Answer 3

1) Denaturation
2) Annealing
3) Elongation
4) Steps 1-3 are repeated

Question 4

Denaturation

Answer 4

DNA is heated to break hydrogen bonds/separate it into single strands

Question 5

Annealing

Answer 5

DNA primers attach to the 5’ ends of target sequence at low temps

Question 6

Elongation

Answer 6

Taq DNA polymerase adds nucleotides

Question 7

What do primers allow?

Answer 7

the selective copying of a DNA region, bind on either side of the target sequence (on opposite strands)

Question 8

Taq DNA Polymerase

Answer 8

heat tolerant enzyme

Question 9

Restriction endonucleases

Answer 9

enzymes that cut DNA into fragments based on recognition of certain sequences

Question 10

What do restriction endonucleases in DNA?

Answer 10

cut sugar-phosphate

Question 11

What does an restriction endonucleases generate?

Answer 11

sticky or blunt ends

Question 12

Sticky ends are…

Answer 12

complementary and single stranded

Question 13

Gel Electrophoresis

Answer 13

A laboratory technique used to separate and isolate protein or DNA fragments based on size

Question 14

How are fragments in gel electrophoresis made?

Answer 14

by restriction endonucleases

Question 15

How does gel electrophoresis work (5)?

Answer 15

• Samples of DNA or protein are placed in a porous gel
• Placed in a chamber and covered with a salt solution, conducts electricity
• The chamber has a negative electrode, where DNA is placed, and a positive
• Power supply generates current that moves samples to positive end
• Small samples move faster than large ones through the gel’s small holes

Question 16

Genomics

Answer 16

the study of an organism’s complete set of genes and their interactions

Question 17

Human Genome Project (HGP)

Answer 17

an international effort to determine the complete sequence of DNA bases in the human genome

Question 18

What were the results of the HGP?

Answer 18

• Humans have 21,000 genes in 3.2 billion nucleotide pairs
• Only 1-2% of the DNA codes for proteins (genes)

Question 19

DNA Sequencing

Answer 19

process by which the nitrogenous base order of a nucleotide sequence is explained

Question 20

What does DNA sequencing use?

Answer 20

• PCR (with dideoxynucleotides)
• electrophoresis

Question 21

Dideoxynucleotides (ddNTPs)

Answer 21

nucleotides that lack the 3’-hydroxyl group necessary for forming phosphodiester bonds; terminate replication

Question 22

What is the difference between regular PCR and PCR for DNA sequencing?

Answer 22

fluorescently labeled dideoxyribonucleotides are included with normal nucleotides in the PCR solutions

Question 23

What is different about DNA strands coming from PCRs using ddNTPs?

Answer 23

they differ in length

Question 24

What does the length of a DNA strand indicate?

Answer 24

the position where a ddNTP was incorporated

Question 25

What does reading the dideoxyribonucleotides from the shortest to longest fragment do?

Answer 25

it gives the base sequence of the original molecule in the 3’ to 5’ direction

Question 26

How are fluorescently labeled DNA fragments separated?

Answer 26

based on size using gel electrophoresis

Question 27

What happens as each DNA fragment reaches the end of the gel?

Answer 27

a laser excites its fluorescent base and a camera detects the color and therefore the base

Question 28

DNA profiling (2)

Answer 28

- the analysis of DNA fragments to determine whether they come from an individual - compares sequences from noncoding DNA regions that show variation between individuals

Question 29

Where is DNA profiling used?

Answer 29

paternity and forensic investigations

Question 30

Why aren't genes used in DNA profiling?

Answer 30

they are too similar between individuals

Question 31

short tandem repeats (STRs)

Answer 31

long stretches of DNA in non-coding regions made up of repeating elements

Question 32

Who founded the technique of DNA sequencing?

Answer 32

Fred Sanger

Question 33

Why are STRs useful (2)?

Answer 33

- Every person has several locations were STR repeats are present in their DNA (same locations across different people) - Each person has a unique number of STR repeats at each location in their genome

Question 34

How can STRs be cut out?

Answer 34

using restriction endonucleases

Question 35

What about STR fragments help with identification?

Answer 35

their length

Question 36

What is needed for there to be a match between crime scene and suspect DNA?

Answer 36

DNA fragments of suspects should be a complete match with the DNA fragments from crime scene

Question 37

What are the steps of DNA profiling (4)?

Answer 37

- Polymerase chain reaction (PCR) is used to amplify a DNA sample collected from an individual - Restriction endonucleases are used to cut out short tandem repeats from DNA - Gel electrophoresis separates the fragments of DNA - DNA fragments are analyzed

Question 38

How are DNA fragments analyzed for forensic investigations?

Answer 38

DNA of suspect compared to DNA from a crime scene

Question 39

How are DNA fragments analyzed for paternity investigations?

Answer 39

child’s DNA fragments are compared to its suspected parents

Question 40

What must be seen for someone to be considered the biological father of a child on a paternity test?

Answer 40

The DNA that is not covered by the mother must be made up for by the father