Equipment and Materials

Week 1

• Previously harvested frozen grape leaves
• Qiagen DNeasy Plant Mini Kit (50) (Qiagen, Valencia, CA)
• 70% EtOH
• Liquid N₂
• Mortar and pestle
• Homogenization pestle for 1.5 ml microcentrifuge tubes
• 0.2 ml PCR tubes
• Degenerate PCR primers
  • LM 638 (5′-GGIGGIGTIGGIAAIACIAC-3′)
  • LM 637 (5′-A(A/G)IGCTA(A/G)IGGIA(A/G)ICC-3′)
• NanoDrop™ One microvolume UV-Vis spectrophotometer (Thermo Scientific, Wilmington, DE)
• T-100 Thermal Cycler (BioRad USA, Hercules, CA)
• Promega 2X PCR Master Mix (Promega US, Madison, WI)
• JumpStart Taq DNA polymerase (MilliporeSigma, Burlington, MA)

Week 2

• 1000X GelRed (Phenix Research Products, Candler, NC)
• 50X TAE buffer
• Agarose
• Power supply
• Ice bucket
• QIAquick PCR Purification Kit (Qiagen, Valencia, CA)
• Agarose gel electrophoresis apparatus
• Biospectrum UVP GelDoc-It®2 Imager (Analytik Jena, Upland, CA)

Week 3

• Pre-cut parafilm strips
• 37°C incubator
• Ice bucket
• L-shape spreader
• 42°C water bath
• 37°C shaker

LB Agar (per Liter)

• 10 g of NaCl; 10 g of tryptone; 5 g of yeast extract; 20 g of agar
• Add deionized H₂O to a final volume of 1 liter.
• Autoclave, pour into Petri dishes (~25 ml/100 mm plate)

LB Broth (per Liter)

• 10 g of NaCl, 10 g of tryptone, 5 g of yeast extract
• Add deionized H₂O to 1 liter
• Autoclave

LB-Ampicillin Agar (per Liter)

• 1 liter of LB agar, autoclaved, cool to 55°C
• Add 10 ml of 10 mg/ml filter-sterilized ampicillin
• Pour into Petri dishes (~25 ml/100 mm plate)

LB-Kanamycin Agar (per Liter)

• 1 liter of LB agar, autoclaved
• Cool to 55°C; add 2.5 ml of 20 mg/ml filter-sterilized kanamycin
• Pour into Petri dishes (~25 ml/100 mm plate)

2% X-gal (per 10 ml)

• 0.2 g of 5-Bromo-4-chloro-3-indolyl-β-Dgalactopyranoside (X-gal)
• 10 ml of dimethylformamide (DMF)
• Store at -20°C. Spread 40 µl per LB agar plate.