Category Archives: Protocols

Reaction   EX: For a 20 ul total volume of reaction (15 ug DNA)                   5 ul DNA: Maximum DNA 1/3 of total volume          2 ul Buffer 10X: Maximum 1/10 of total volume          2 ul enzyme: NEB … Continue reading →

Digest good-quality genomic or plasmid DNA to completion with restriction enzymes. Use AT LEAST 1 ug  of genomic DNA per lane; 10 micrograms per lane is ideal.  If more than one DNA sample is to be compared, you should load … Continue reading →

Protocol used to concentrate poly(A)+ mRNA, RNA, DNA. Also to clean reactions from buffers and others that may interfere in dowstream reactions.   1. Add 1/10 volume 3 M NaAc (Sodium acetate) and 3 volumes of cool ethanol 95% (high … Continue reading →

  Vectors contain sequences that allow plasmid recovery (rescue) including genomic sequences from the transformed fungus. Sequences required for plasmid rescue are a bacterial origin of replication and an antibiotic resistance gene (e.g. Ampicillin or Kanamycin resistance). In addition, unique … Continue reading →

THE BEST METHOD WE HAVE FOUND (modify from Lisa Vaillancourt Lab) (Method of John Tuite, modified by Rick Latin, 9-1-95) 1. Fill vials  (cryovials Wheaton 8 ml, cat 224884 we get them from VWR) half full with silica gel granules, … Continue reading →

(from Slavica)   5 minutes 70% ethanol. 1:45 hours 10% H2O2   In a 50 mL tube, rinse seeds with sterile water 4-5 times.   If seeds will be used for In vitro tissue culture: Seeds will float on H2O2, … Continue reading →

                                                                                                                  (Modification from Lisa Vaillancourt Lab)    1. We use TOP10F’ cells (Electrocomp Kits, INVITROGEN cat C6665-55, Is in the -80C. READ CAREFULL Kit information, print … Continue reading →

  1. Harvest conidia from 2 or more plates that are 3-4 weeks old, center inoculated. Expect about 1×108 conidia per plate. 2. Wash conidia once with sterile water. Resuspend the last time in Fries medium. 3. Count with hemacytometer. … Continue reading →

1. Colletotrichum graminicola is grown on Petri dishes of Potato Dextrose Agar (PDA) at room temperature under fluorescent lights. The cultures should be 12-15 days old. 2. Place 8-10 mL of sterile distilled water on the surface of the Petri … Continue reading →

From Lisa Vaillancourt Lab   1. Prepare PEG solution and place on a rocker to mix. 2. Thaw protoplasts on ice. 3. Place 100 mL (10^7 protoplasts) in a pre chilled 50 mL centrifuge tube. 4. Add 3 ug of … Continue reading →