6. stkP reverse cloning primer: TAAGCAGCGGCCGCAGGA
GTAGCTGAAGTTGTTTTA.
7. dexB forward pLF primer: GCGGGATCCCAAGAAAAAT
GGTGGCATAATGCCGTAG.
8. dexB reverse pLF primer: ATAGGCGCGCCTTATAGTAA
TTCCACACAG.
9. glpO forward pLF primer: GCGGTCGACAAGGAGATATAA
TGGAATTTTCAAAAAAAAC.
10. glpO reverse pLF primer: GCGGCGGCCGCTTAATTTTTT
AATTCTGC.
11. stkP forward pLF primer: GCGCATATGATCCAAATCGGC
AAGATTTTTG.
12. stkP reverse pLF primer: GCGCAATTGTTAAGGAGTAGC
TGAAGTTGTTTTAG.
13. pspA forward pLF primer: ATAGGCCGGCCAAGGAGATAT
AATGGAAGAATCTCCCGTAGCCA.
14. pspA reverse pLF primer: ATACTCGAGTTATTCTGGGGCT
GGAGTTTCTGGA.
15. pUAB055 containing pspA.
16. Phusion High-Fidelity DNA polymerase.
17. XbaI, AgeI, XhoI, NdeI, SacI, NotI, BamHI, AscI, SalI, MunI,
and FseI restriction enzymes.
18. Plasmid Miniprep kit.
19. GeneJET Gel Extraction Kit.
20. Agarose.
21. T4 DNA ligase.
2.2 Cell Culture
Materials
1. Todd Hewitt blood agar plates: 10 g/L pancreatic digest of
casein, 60 g/L yeast extract, 3.1 g/L beef heart infusion,
2.5 g/L sodium carbonate, 2 g/L dextrose, 2 g/L sodium
chloride, 0.4 g/L sodium phosphate, and 5% v/v sheep blood.
2. Lysogeny broth (LB) medium: 10 g Bacto tryptone, 5 g yeast
extract, 10 g/L NaCl, and antibiotic (100 mg/L ampicillin,
50 mg/L kanamycin, or 20 mg/L chloramphenicol, as
needed).
3. LB agar plates: 10 g/L Bacto tryptone, 5 g/L yeast extract,
10 g /L NaCl, 15 g/L agar, and antibiotic (100 mg/L ampicillin, 50 mg/L kanamycin, or 20 mg/L chloramphenicol, as
needed).
4. RPMI-1640 cell culture media: 10% FBS, 5 mM sodium pyruvate, 1% penicillin/streptavidin, 25 mM HEPES.
5. THY Media: 37 g/L Todd Hewitt Broth, 0.5% yeast extract.
Hybrid Biological-Biomaterial Vector
463