D1.1- DNA replication

Question 1

why DNA replication is required

Answer 1

organisms must replicate their entire genome for; -growth, -replacement of damaged tissue, -reproduction

Question 2

why DNA replication is semi-conservative

Answer 2

DNA of parent must be separated and bonds broken, each strand is used as a template for the assembly of a new polymer. One new strand is conserved from the parent, and one is newly replicated which are identical to template strand

Question 3

role of helicase and DNA replicase

Answer 3

helicase= enzyme which unwinds the double helix and separates the two strands by breaking the H bonds between bases
DNA replicase= adds nucleotides to the 3’ end of a primer, building the new strand

Question 4

steps and procedure of polymerase chain reaction

Answer 4

PCR= used to copy DNA and amplify targeted section
components needed:
1) DNA sample
2) taq polymerase (polymerase from bacteria in hot springs, which contains enzymes that withstand high temps)
3) Primers (short polymers with the complementary nucleotides to targeted DNA. This binds to DNA after separation
4)Free nucleotides (to synthesize new DNA strands

STEPS:

1) DENATURATION: DNA is heated to 95* to break H bonds between strands

2) Annealing: Mixture is cooled to 60* to allow primers to bind (anneal) to nucleotide on both strands of targeted DNA

3) elongation: temp increases to 72*, optimum temp for taq polymerase, to build complementary strands for DNA

This cycle is repeated

THEN GEL ELECTROPHORISIS

Question 5

Gel electrophoresis

Answer 5

mixture is placed into small wells and agrose gel is added. The mixture is exposed to an electric current. positive on one side and negative on the other. Largest, heaviest and least charged particles dont move far, the smallest, most charge particles pass through the gel and move far.

Question 6

application of PCR and gel electrophoresis

Answer 6

1) forensic science: evidence in court cases, DNA profile matching DNA in crime scenes.

2) Paternity test: DNA profile of child and mother. All bands are compared with mother and true father.

Question 7

Directionality of DNA polymerase

Answer 7

DNA polymerase adds the 5’ of a nucleotide to a 3’ end of a strand of nucleotides. The stands are antiparallel and therefor replication works differently on each strand.

Question 8

replication on the leading and lagging strand

Answer 8

Leading= RNA primer is assembled at the start of the leading strand. DNA polymerase III moves in the same direction as the replication works as the replication fork. Replication is continuous.

Lagging: DNA polymerase III moves away from the replication fork, adding nucleotides to the growing chain until it reaches the previous RNA primer (5’-3’). These series of short lengths of DNA are assembled on the lagging strand, called okazaki fragments. Replication is discontinous

Question 9

function of enzymes in replication

Answer 9

helicase= unzips double helix at replication fork

DNA primase= synthesizes an RNA primer

RNA primer= marks the beginning of replication and replaces it with DNA nucleotides

DNA polymerase I= Removes RNA primer at the end of replication and replaces it with DNA nucleotides

DNA polymerase III= snythesizes new DNA strands, by adding nucleotides in a 5’-3’ direction

DNA ligase= joins the end of the DNA segments and okazaki fragments

Question 10

DNA proofreading

Answer 10

DNA polymerase III adds nucleotides according to complementary base pairing. When there is a mismatch base, it removes from from the 3’ end, and replaces it, then reverses the direction and resumes synthesis; reducing mutations in replication