IOWA STATE UNIVERSITY | DEPARTMENT OF AGRONOMY | COLLEGE OF AGRICULTURE AND LIFE SCIENCES | PLANT SCIENCES INSTITUTE

1. Submit  2. APHIS   3. Analysis   4. Plantlets or Seed

Submit your construct- Biolistic

  1. Purify your plasmid DNA using a Qiagen plasmid DNA preparation kit. Dissolve DNA in ddH2O, not TE!
  2. Measure DNA concentration using a spectrophotometer. Obtain OD260 and OD280 readings. Dilute DNA to 1 µg/µl concentration. An acceptable DNA purity is within an OD260/280 range of 1.6 to 1.9.
  3. Check DNA quality (1 µg/lane) using electrophoresis by including
    1. undigested DNA
    2. DNA digested with 2-3 restriction enzymes to confirm the construct.

      Restriction digestion

  4. Ship DNA to PTF via overnight express (ice is not needed). The package should include:

    1. Construct map

      10-20 µg DNA in ddH2O (concentration of 1µg/µl), packaged as indicated in #5.
    2. Brief physical map of the construct with
      1. major restriction enzymes positions
      2. size of the construct
    3. OD260 and OD280 values and ratio
    4. Gel picture illustrating the DNA quality (as described in #3).
    5. Title and brief description of your project including number of events desired and type of product (seed, plant, callus).
    6. Your complete contact information including the name of PI, name of contact person for construct communications and receiving samples, lab and office phone numbers, fax number, and e-mail addresses.
  5. Plasmid DNA must be packaged as follows:
    1. Plasmid DNA must be in a primary inner container (1.5 ml tube)
    2. Secure tube lid with parafilm (see below)
    3. Place tube in a sealable secondary container (plastic bag)
    4. Place a towel or kimwipe in the bag and seal (see below)
    5. Place bag in a cushioned envelope or box (see below)
    6. Enclose an itemized list of package contents
    7. Seal and mail

  6. You will be notified by an e-mail message of the Identification Number (ID#) that has been assigned to your construct. PTF personnel will thereafter communicate with you using this ID# rather than the original name of your construct.