Identifying a Body

Question 1

What is a DNA primer? What are they marked with?

Answer 1

• short DNA sequences complementary to the DNA adjacent to the STR
• primers are marked with fluorescent tags

Question 2

What is a nucleotide?

Answer 2

• a nucleotide is an organic molecule with a basic composition of a nitrogenous base, pentose sugar and phosphate
• it is the basic building block of nucleic acids (RNA and DNA)

Question 3

What is DNA polymerase?

Answer 3

• DNA polymerase is an enzyme that synthesises DNA by adding nucleotides one by one to the growing DNA chain

Question 4

What does the process annealing refer to?

Answer 4

• annealing is the process of joining of single-stranded DNA or RNA by hydrogen bonds to form a double-stranded polynucleotide
• when two complementary strands of DNA or RNA have paired with each other or a synthetic primer

Question 5

What does PCR stand for? What is used to make?

Answer 5

• PCR stands for polymerase chain reaction
• polymerase chain reaction is used to make (numerous) new copies of all the fragments of DNA present (‘amplifying’ the DNA)
• often only small amounts of DNA are available for forensic analysis so the DNA fragments loci are copied to get enough DNA to make a profile

Question 6

What does STR stand for? What are STR’s? How many base pairs does it contain and how many times can it be repeated?

Answer 6

• STR stands for short tandem repeats or satellites
• STR’s are sections of repetitive DNA where a short sequence is repeated multiple times (STR’s are sequences or repeated bases)
• An STR can contain from 2 to 50 base pairs and can be repeated from five to several hundred times

Question 7

What are introns? Where are they found?

Answer 7

• introns are the non-coding regions of the gene that do not contain codons needed to make the final protein (intragenic regions)
• introns are found between exons

Question 8

What are exons?

Answer 8

• exons are the coding regions of the gene (expressed regions)
• they contain the codons that are later read to make protons

Question 9

What is southern blotting used to do? What is it named after?

Answer 9

• southern blotting (named after Ed Southern who developed the technique) is used to transfer the fragments to a more resilient nylon or nitrocellulose membrane

Question 10

What happens to the fragments during southern blotting?

Answer 10

• during southern blotting process the fragments maintain their positions relative to each other and are denatured into single strands exposing base sequences
• DNA denatures in alkali solution

Question 11

What is a DNA probe? What can the probes be labelled with and why?

Answer 11

• a DNA probe is a short, single-stranded section of DNA that is complementary to the target DNA sequence
• the DNA probe can be labelled with a radioactive marker or a fluorescent marker to help see the location of the DNA fragments

Question 12

What is a DNA ladder/ marker added to?

Answer 12

• a DNA ladder or marker is sometimes added to the gel (in gel electrophoresis)

Question 13

What does gel electrophoresis separate? What does this depend on?

Answer 13

• DNA fragments produced by restriction enzymes or PCR can be separated by gel electrophoresis depending on their size

Question 14

What are three ways in which a body can be identified?

Answer 14

a body can be identified by personal identification, fingerprinting or dental records

Question 15

What are three ways in which a DNA sample can be obtained?

Answer 15

• cells obtained in a cheek swab
• white blood cells in a blood smear
• bone marrow in a skeleton
• sperm left after a sexual assault

Question 16

How is DNA extracted (think back to kiwi)?

Answer 16

• tissue sample is physically broken down in a buffer solution including salt and a detergent to disrupt the cell membranes
• small suspended particles, including DNA, are separated from the rest of the cell debris by filtering or centrifuging
• protease enzymes are incubated with the suspension to remove proteins
• cold ethanol is added to precipitate out the DNA
• several stages of then washing the DNA in a buffer solution then follow

Question 17

How is a DNA profile made?

Answer 17

1. obtaining the DNA
2. Cut the DNA into fragments using restriction enzymes
3. Amplifying DNA present in the sample (polymerase chain reaction)
4. separate out the fragments into a characteristic pattern (DNA fingerprint) using gel electrophoresis
5. southern blotting to transfer DNA onto membrane
6. disclosure - show the presence of the DNA using a DNA probe or staining. Loading dye can be used to show progress of DNA through gel

Question 18

What are restriction enzymes? What are they used for in labs?

Answer 18

• restriction enzymes (aka restriction endonucleases) are bacteria produced enzymes that are used to cut DNA into smaller fragments only where their specific restriction sequences occurs
• they are used as targeted ‘scissors’ in the lab

Question 19

How is a DNA profile made from a small sample of DNA? (in own words)

Answer 19

• multiple copies of DNA made using polymerase chain reaction (PCR)
• step 1: 90 degrees to 95 degrees
  step 2: 50 degrees to 65 degrees
  step 3 70 degrees to 80 degrees
• restriction enzymes used to cut the DNA into smaller fragments
• gel electrophoresis used to separate the fragments
• DNA is placed onto a gel of agarose
• an electric current is applied
• southern blotting is used to transfer DNA onto membrane
• the presence of DNA is shown using a fluorescent tag

Question 20

How can DNA profiles be compared?

Answer 20

• DNA profiles can be compared by comparing the total number of bands
• comparing the position of the bands
• comparing the size/ width of the bands