Topic 7 Flashcards
Pyrimidines
CT
Purines
AG
What does C pair with
G
What does A pair with
T
A+G=
T+C
A=
G=
T
C
What does every full twist of a dna have
10 base pairs (5 paired with 5)
How many h bonds does gc have
3
What is the 3’ end
The one with the oh
What carbon of the sugar is the phosphate bonded to
The nucleotide
5’ and 3’ carbons
1’ carbon
What is cleaved off to make the rest of the replicated dna strand
Pyrophosphate, give NRG to drive the rxn
New stuff is added to which end
3’
Synthesis in 5-3
What does helicase do
Break h bonds between nucleotides In The dna strand
What do ss dna binding protiens do
Stabilize the unwound dna from helicase
What does gyrase do
Type of topoisomerase
Relaxes the dna strands and stops them from coiling up and then rejoins the strands. It’s behind the replication fork
What does DNA pol. III do
Catalyzed dna synthesis. Extends existing dna strand but cannot start a new one
5-3
Leading stand and laggin strand
Leading strand is the strand synthesized from the top 5-3 template strand
The laggin strand is the strand made from the bottom 3-5 template
Dna pol 1
Removes the rna primer and fills gap left behind
Dna ligase
Connects the Okazaki fragments
Rna polymerase
Makes the rna primer which is the starting pint for dna pol
Dna ligase makes what type of bond
Phosphodiester bond
What is PCr (polymerase chain rxn)
A way to amplify a piece of dna (double helix)
Make a billion of the same dna
What does you need for PCr
- Template dna: Extract all dna (genomic dna) and analyze small portion of a gene in the dna
- Design dna primers that have a free oh group that bind to the region of the gene you’re looking at
Need a forwards primer (complementary to bottom template strand) need reverse primer( reverse complement to top template strand) - Need dntps (agtc)
- Tac polymerase: type of dna pol to synthesis the dna, best polymerase for PCr
What happens in PCr
- Heating at 95 degrees, denatures h bond and separates the two strands
- Cooling and 55-66 degrees. Anneals the primers to their location on the template dna
3.heat to 72 degrees, synthesis, taq pol adds to 3’ end of Top and bottom primers
All these step repeated over and over to amplify and make billion copies
What determines the corrects annealing temp in PCr
The melting temp of the primer
The melting temp is the temp where 50% of primers are are bound to dna and 50% aren’t
What determines the melting temp of the primer
The amount of gc
More gc higher melting temp
Length of the primer, longer higher melting temp
The aneealing temp should be ______ than the melting temp
Lower
To encourage binding
What is gel electrophoresis
A method to separate the dna molecules based on their size. Use el tric field to move the dna through the gel
To check if PCr amplified a region of the the size we wanted
Why is gel electrophoresis negative and positive charge
Since dna is negative charge, they will travel towards postive charge at the bottom of the gel.
Negative charge is at the wells
Why do we stain the dna in gel electrophoresis
To see the dna in the gel, dna is colourless so we add colour the fluorescence under uv
What is the molecular ladder
Had dna prices of known sizes that determin size of the PCr dna
Why do we use taq pol in PCr
Active at high temps and remains active
