Protocols

Ligation Protocol

Determine insert to vector ratios

Calculate the amount of insert needed if 50ng of vector is used (can use different amount of vector)

In a PCR tube add the following:

50ng of vector

Amount of insert based on ratios (calculated in second step)

2uL of buffer

2uL of DNA ligase

Amount of water to bring total volume to 20uL

Incubate overnight at 14oC

Note: We used T4 DNA ligase and buffer from NEB

Gel Purification Protocol (QIAquick Gel Extraction Kit)

Gel Electrophoresis Protocol

Digestion Protocol

Preparing LB+Appropriate Antibiotic Protocol

Preparing Agar Plates Protocol (Makes 12 (15mm) Plates)

Preparing Competent Cells Protocol

Miniprep Protocol (from QIAprep Spin Miniprep Kit)

Preparing Glycerol Stock Protocol

Transformation Protocol