Instructions: Cover the answers and fill in the blanks. Then check.

1. The recognition sequence of EcoRI is _______ and it produces _______ ends. Answer: 5'-GAATTC-3'; sticky ends

2. Taq DNA polymerase is isolated from _______, found in _______. Answer: Thermus aquaticus; hot springs

3. DNA migrates toward the _______ (_______ electrode) during gel electrophoresis. Answer: anode; positive

4. pBR322 has two selectable markers: _______ resistance and _______ resistance. Answer: ampicillin (ampR); tetracycline (tetR)

5. In PCR, the three temperature steps are: _______ at ___°C, _______ at ___°C, and _______ at ___°C. Answer: Denaturation at 94-98°C; Annealing at 50-65°C; Extension at 72°C

6. After n cycles of PCR starting from 1 molecule, _______ copies are produced. Answer: 2^n

7. DNA bands in agarose gel are stained with _______ and viewed under _______ light. Answer: ethidium bromide; UV (ultraviolet)

8. The Ti plasmid is from _______ and is used for _______ transformation. Answer: Agrobacterium tumefaciens; plant

9. The _______ in a stirred-tank bioreactor provides sterile air to the culture. Answer: sparger

10. During DNA isolation, bacterial cells are lysed by _______, while plant cells are lysed by _______. Answer: lysozyme; cellulase

11. Competent E. coli cells are prepared by _______ treatment followed by _______ at 42°C. Answer: CaCl2; heat shock

12. PCR was developed by _______, who won the Nobel Prize in Chemistry in _______. Answer: Kary Mullis; 1993

13. DNA fired on gold or tungsten particles into cells describes the _______ transformation method. Answer: biolistics (gene gun)

14. Recombinants in pBR322 cloning grow on _______ but do NOT grow on _______. Answer: ampicillin; tetracycline

15. The downstream processing step that immediately follows bioreactor production is _______. Answer: separation $\frac{of cells}{debris from medium}$

16. Over _______ restriction enzymes have been found from over _______ bacterial strains. Answer: 900; 230

17. Restriction enzymes recognize _______ sequences and make cuts that produce _______ or _______ ends. Answer: palindromic; sticky; blunt

18. The _______ gene in the Ti plasmid is retained in the disarmed version because it mediates T-DNA transfer. Answer: vir (virulence) genes