Prepared February 8, 1996                   Heavenly Gardens
SOP No.  001
by James M. Gossard                                Standard Operating
Procedure
James Gossard      1069 Amity  Road,  Galloway , Ohio 43119-9304     (614)
853-0522        e-mail    JGOSSARD@AOL.COM

Surflan Seed Conversion Method
                        Kevin Vaughn Method USDA
A.  Reagents

              Note: ( Label all reagents with name, conc., date made, expiration date, and storage temperature. )
   (1)  Preparation of 10(M Oryzalin solution

    (i)   Oryzalin -99.9% pure     (Molecular Weight - 346.36)
                  Also known as  3,5-dinitro-N4N4-dipropyl-sulfanilamide.
                  Pure reagent available from Elanco Products Co., Division of Eli Lilly & Co.

    (ii)  Stock Solution - Using 99.9% pure Oryzalin make a 10 (M solution in  DMSO.
           Dissolve 0.3463 grams Oryzalin in DMSO  ( Dimethyl Sulfoxide ) and dilute to 100 mls with DMSO.

    (iii) Working solution  -10 (M (10-5 M solution  =  3.4636 mg/L)
           Dilute 1 ml of stock solution to 100 mls in Distilled or D.I. Water. ( 1-2 mls DMSO may be added if desired.)

   (2)  Preparation of 10(M Oryzalin solution using Surflan AS

    (i)   Surflan AS - sold by Dow-Elanco as a pre-emergence herbicide under the name Surflan AS.
                  Purity of oryzalin in  surflan AS is 40.4% , thus stock solution should be adjusted accordingly.

    (ii)  Stock Solution - Using 40.4% purity Oryzalin make a 10 (M solution in Distilled or D.I. Water.
           Dissolve 0.857 grams ( approx. 1 ml)  Surflan AS in Distilled or D.I. Water and dilute to 100 mls.

    (iii) Working solution  -10 (M (10-5 M solution  =  3.4636 mg/L)
           Dilute 1 ml of stock solution to 100 mls in Distilled or D.I. Water. ( 1-2 mls DMSO may be added if desired.)

B. Procedure

          1) Germinate seed in a container with wet filter paper or paper towels.  An alternative method under less
              sterile conditions would be to germinate seeds on Pro-Mix or equivalent.

          2) Germinate seeds under lights - 24 hours on.

          3) Treat seeds 5-8 days from start of germination - seeds must show germination.

          4) Treat germinated seeds for 24 hours in 10 (M working solution of Oryzalin in the dark. Use glass
              containers or glass petri dishes, plastic may absorb the oryzalin.  Also do not use cheesecloth, as it may
              also absorb the oryzalin.

          5) After 24 hours the seedling roots and the base of the growing shoot should be swollen, as an indication
              that treatment was effective.

          6) Rinse seeds wrapped in cheesecloth or pantyhose  for 4 hours under slowly running water.

          7) Transfer to promix or similar growing medium.. Plant  at surface of mix, but not too deep as seed is
              prone to rot. Cover with plastic bag to prevent moisture loss.

          8) A cool 72 F dimly-lighted area ( 1/10 th full sunlight area ) works well until good growth occurs.