D1.1 DNA Replication

How is new DNA produced? How has knowledge of DNA replication enabled applications in biotechnology? How is biotechnology unique in terms of human ingenuity? What biotechnology processes have been developed for human benefit? How old is biotechnology?

what is semi-conservative DNA rep

•Each DNA molecule formed by replication consists

-one new strand

-one old strand conserved from the parent DNA molecule. 

•The result of replication is therefore is two DNA molecules, both composed of on original strand and a newly synthesized strand.

what three functions need DNA rep?

reproduction, growth, and repair

what did the meselson stahl experiment show?

•The two DNA strands separate and each original strand serves as a template for the creation of the new strands.

(semi conservative rep)

what does helicase do

-Unwinds DNA helix by breaking H-bonds

what is helicase made of

•Globular protein composed of 6 polypeptides arranged in a doughnut shape.

In what drection does DNA polymerase I build in?

5’ —> 3’

what are PCRs

(polymerase chain reactions) A method to amplify any piece of DNA

where does Taq DNA polymerase come from?

-Extracted from Thermus aquaticus, which can be found in hot springs.

what is gel electrophoresis?

-Used to visualize separate DNA (RNA, or protein) molecules by size

-Place molecules at one end of a jello-like matrix (agarose) and apply an electric current

•The molecules move along the current towards the positive pole according to size

•Larger pieces of DNA have a harder time squeezing their way through the gel than smaller pieces and therefore migrate a shorter distance

what determines how far molecules move in gel electrophoresis?

molecule size

Where do VNTRs come from?

inherited from both parents

what did Lnus Pauling conclude about DNA?

concept of molecular modelling

what did rosalind franklin conclude about DNA?

X-ray diffraction

what did Erwin Chargaff conclude about DNA?

base composition

What did Watson and Crick conclude about DNA?

-Tight packing of DNA would occur if a pyrimidine was paired with a purine.

-Bases were “upside down” (antiparallel). 

•This suggested the mechanism for semi-conservative replication.

-Stability

how many bonds do CG have?

3 hydrogen bonds

how many bonds do AT have?

2 hydrogen bonds

where does the energy to form phosphodiester bonds come from?

the hydrolysis of the first 2 phosphates from the nucleotide

what kind of reaction is the addition of new nucleotides?

dehydration synthesis/condensation

how many primers does the leading strand need?

1

how many primers does the lagging strand need

>1

ligase function

Connects gaps between Okazaki fragments

topoisomerase function

Releases the supercoiling strain that develops ahead of helicase

DNA primase function

Creates an RNA primer (~20bp) and initiates the activity of DNA polymerase.

One primer on leading strand, many primers on lagging strand.

DNA polymerase III function

Adds complimentary nucleotides to growing chain

Proofreads

DNA Polymerase I function

Removes RNA primers, both an exonuclease (it breaks phosphodiester bonds) and polymerase

what happens when pol III notices a error in replication?

When a disturbance in the normal helical shape is discovered, pol III removes the mismatch from the 3’ end and replaces it with the correct base

how many origins of rep do prokaryotes have?

one origin of replication but is bidirectional

how many origins of rep do eukaryotes have?

multiple origins of replication (makes process faster)

What enzyme catalyzes the elongation of the leading strand?

DNA polymerase

how often will the restriction enzyme EcoR cut DNA equences.

randomly

Which carbon of the ribose (pentose) sugar attaches to the phosphate in a nucleic acid?

C5