实验动物模型

N2 development times at different temperaturesby Michael KoelleDetermined empirically; times in hours are given from the first division of the zygote.15°20°25°eggs laid217hatch22139L1 molt492921L2 mo ...

low frequency tableCodon usage for C. elegans lowly biased (lowly expressed by inference) genes. (Stenico M. Lloyd A. T. and Sharp P. M. (1994). Nucleic Acids Research 22: 2437-2246 AmAcidCodonNumber/ ...

Worm genomic Southern blotsby Michael Koelle4/6/94I. Preparing worm genomic DNA: requires 1-2 days to seed agarose plates a few days for the worms to grow 1-2 days to prep the DNA1. Seed large agarose ...

Worm PCRPick one worm and place it in a 2.5 l drop of lysis buffer in the cap of a PCR tube. Close and centrifuge briefly to move to the bottom of the tube. Freeze the tubes at -70°C for 15 min. The ...

Genomic DNA prep(Jorgensen Lab ) Make worm growth plates. These use agarose to avoid impurities in most batches of agar and are enriched to allow greater worm growth. Mix:5 g Bacto tryptone2 g Bacto y ...

Joe's mRNA prep(Audrey Gasch Pat Brown ) Oligo-dT cellulose prepDump Ambion vial contents into 50 ml c/f tube Add 10 ml 1x NETS to vial cap rinse vial dump 10 ml into c/f tube Spin tube 3000 rpm 2 min ...

Yale RNA prep(Rebecca D. Burdine Michael J. Stern ) Add 8 ml of TRIZOL to 2 ml packed worms in 15 ml centrifuge tube. Vortex and invert tube to solubilize and lyse worms for at least 10 min. Divide in ...

Antibody Staining of Dissected Gonads - From R. Francis - -Adapted by Min-Ho Lee-Antibody staining (in Tubes) 1. Using a pasteur pipette (drawn in flame and cut open) transfer worm carcasses with a ...

RUVKUN Antibody Staining Protocol revised 10/29/90Fixation: 1. C. elegans N2 grown in liquid or plates. Harvest worms.Wash for 2 hours at room temp. in 1X M9 or PBS. 2. Fix in 5 ml of FRESH THAT DAY 1 ...

C. elegans whole mount immunohistochemistry with Bouin's FixativeI have developed a fixation protocol which provides an alternative set of conditions to test antisera that work poorly on paraformaldeh ...

DAPI Staining To visualize DNA incubate fixed samples with 100 ng/ml 4'6'-diamidino-2-phenylindole hydrochloride (DAPI) in PBS for 30 min. Rinse 3x with PBTw. MaterialsPBS: Sambrook et al. (Molecular ...

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 ...