1. SURVEY OF BIOCHEMISTRY Nucleic Acids continued… Amino Acids

2. Name the following compound
PRS Question
Name the following compound
ATP
UTP
GTP
CTP

3. Thymine has a methyl group at which position?
PRS Question
Thymine has a methyl group at which position? C1 C3 C5
C1’

4. Recombinant DNA Technology
Last Week…
Recombinant DNA Technology
Vectors, Cloning and Libraries
Genomic vs. cDNA Library
How are libraries made?
PCR (for making copies of DNA)
How does PCR work?
What is required to conduct PCR?

5. Cut Genomic DNA with restriction enzymes Insert fragments into vectors
Genomic Library
Cut Genomic DNA with restriction enzymes
Insert fragments into vectors
Culture cells
Reference: Wikipedia

6. Making cDNA
Image from Biochemistry by Stryer

7. Image from Biochemistry by Stryer
Making cDNA Clones
Image from Biochemistry by Stryer

8. 3 steps for each PCR cycle
Each PCR cycle includes:
A denaturation step (92-96oC) separates the two DNA strands.
A primer annealing step (40-75oC) which is a few degrees below the Tm of the primers.
A primer extension step (72oC) which is the optimal temperature for Taq DNA polymerase activity.

9. Heat-stable DNA polymerase Thermus aquaticus Taq polymerase
Components for PCR
KNOW THIS!!
Heat-stable DNA polymerase Thermus aquaticus Taq polymerase
Primers (DNA oligonucleotides)
Deoxynucleotides –dATP, dTTP, dCTP, dGTP
DNA template
Mg++, buffer components, and water