实验动物技术

Double dye filling (using DiO and di-4 ANEPPS)(Krista Williams) Dye Filling Label 15 ml centrifuge (c/f) tubes with strain name. Squirt ~1-2 ml M9 onto plate with a Pasteur pipet. Rinse plate with M9 ...

Dye Filling to Stain Amphid and Phasmid Neurons by Michael Koelle from Beth Sawin 4/6/94 1. Buy DiO from Molecular Probes catalog # D-275. DiO fluorescesces green (use FITC filters). If you want red f ...

Fixation and permeabilization protocol 5/30/91 Mike FinneyThis protocol works for all stages except dauers (which won't open) andhypoclorite-treated eggs (which disintegrate). Luckily hypochloritetrea ...

Gonad Dissections- From R. Francis -1) Pick adults and/or L4s to an unseeded NGM plate. Alternatively wash worms off a seeded plate with 1.5 ml of PBS and allow to gravity settle for a few minutes in ...

Gonad Fixation - From R. Francis -A. Methanol Methanol fixation is quick easy and gives satisfactory nuclear morphology after DAPI staining. However methanol-fixed gonads break easily and are more dif ...

Gentle hypochlorite treatment1. Wash off plates of gravid hermaphrodites with 10-15 mls of M9 and put in 15 ml conical tube. Collect minimum of 4 or 5 plates in a tube.2. Centrifuge @500rpm for about ...

In situ hybridizationIn situ hybridizations were based on a modified method of Seydoux and Fire (1994 Development 120 2823-2834; 1995 Methods in Cell Biology) (H. Tabara and Y. Kohara personal commun ...

MMS integration (both methods modified by Herman Lab) NOTES: Remember that MMS is a potent mutagen -- wear gloves and be very careful. Integration is best done using an array that is transmitted at ve ...

Nematode dye filling(Ed Hedgecock ) Thumb plate Transfer worms into a known amount of M9 in a labeled depression. Add DiO(2 mg/ml in DMF) at 10 l/ml of M9. Let sit for 2 hours. Transfer worms with a P ...

RNA In Situ Hybridization of Dissected Gonads with Digoxygenin-Labeled Probes- From R. Francis �- Revised by Min-Ho Lee 2003 -In contrast with other methods using gonads affixed to slide ...

Whole-mount in situ hybridization for the detection of RNA in C. elegans embryosGeraldine Seydoux and Andrew Fire Adapted from Seydoux G. and Fire A. (1995). Whole-mount in situ hybridization for the ...

Freeze-Crack P granule Staining Protocol1. Prepare humidity chamber: line large square plate with humid paper towels and place 4 5ml plastic pipets across chamber to support slides.Place aluminum bloc ...

PAR-1 Staining Procedure Freeze Fracture to permeabilize embryos Fixation 15' in -20 degree 100% MeOH1 x 10' wash in PBSStaining 1 x 60' block in goat serum1 x 60' primary antibody (rabbit-anti-PAR-1 ...

Sos ProtocolAntisera staining/ImmunohistochemistryFixationAMPA assayC. elegans cosmid vectors___________________________________________________________________________________Sos (volatile repellent ...

Phalloidin Staining worms by Michael Koelle modified from Beth Bucher and Andrew Chisholm 4/6/94 Phalloidin binds to filamentous actin. This stain allows visualization of mainly of muscles but some ot ...

Staining C. elegans for ß-galactosidase activity using X-gal by Michael Koelle adapted from Laird Bloom Jeff Way & Michael Basson 4/6/94 1. Grow up a small plate of worms. Some protocols call for usin ...

Protease Inhibitors from Alberts' Lab Protocols (Kathy Miller/Chris Field)Protease inhibitors are rapidly inactivated in aqueous solutions and should be added to solutions immediately before use. Cock ...

Stockkeeping1. MechanicsMost stocks can be successfully cultured by periodic mass transfer of adults to fresh food. Bottles or vials are tapped on the pounding pad to shake flies away from the plug t ...

I. MEDIUMA. S2 S2C Sundefined S2R+ S3 l(2)mbn Kc(167) & DL2 cells: Schneider's/ 10% FBS/ PS450 ml Schneider's medium (Gibco #11720-034)(pour off 50 ml into Falcon to save as serum-free~undefined*50_ml_Fetal_Bovine__...

This is our modification of a recipe used by John Loera at the University of Texas at Austin. The Texas recipe is apparently based on the malt medium described by Lakovaara (1969) with the addition ...