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Note: teaching protocols and materials can be found at "For Teachers: Teaching materials by Fields of Study" under C. elegans education.
Commonly used C. elegans Research protocols
Worm Culture
Dauer
Dauer Pheromone Prep Making dauers with pheromone
Others
Bacteria for worm food “Chunking” worms- easy strain maintenance Cleaning worms by floating on sucrose Cleaning worm stocks Cleaning Worm Stocks Cloning form low melt agarose Cool growth competent E. coli Culturing Worms Decontaminating worm population to remove fungi and bacteria Development of techniques for primary culture of C. elegans Development of techniques for primary culture of C. elegans embryonic neurons Laird Bloom – Part 1 Development of techniques for primary culture of C. elegans embryonic neurons –Part 2 –Part 3 ES cell culture Freezing worms Growing worms in liquid Liquid culture of worms #1 Liquid culture of worms #2 Liquid culture of worms in with fermentor Maintenance of lin2 strain C.elegans Worms N2 Development time at different temperatures #1 N2 development times at different temperatures #2 Preparation pf Petri plates Preparation of synchronized cultures (PDF) Preparation of Seeded NGM Plates For Worm Food Response to food Quick bleach cleaning of small numbers of worms Quick bleach decontamination of small numbers of C. elegans Synchronizing worm cultures Testing for Levasimole resistance
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Solutions
Buffer
Buffers Embryo buffers
Media
S-medium, S-basal and trace metal solution
Mutagenesis
C. elegans gene knockout protocol Chemical deletion mutagenesis DpnI mediated site-directed mutagenesis EMS mutagenesis ems mutagenesis Gene knockout with conventional mutagenesis Irradiation of worms in Y-building facility Making males by heat shock Transposon Insertion Mutagenesis
Mapping
Mapping mutants using STS Polymorphisms Snip-SNP mapping with BSA Snip-SNP mapping with Polymorphisms SNP characteristics
Screening
F2 mutant screen Oligos for screening of deletion libraries Rapid screen of transformants
Injections
Creating Transgenic C. elegans with Extrachromosomal Arrays Integrating extrachromosonal arrays Microinjecting worms Procedures for preparing pads and needles Transformation by injection into syncytial C.elegans germline
Molecular & Biochemical Techniques
Blotting
Western Blot Transfer and Detection Worm genomic southern blots Worm extracts and western blotting
Cell Culture
Development of techniques form primary culture of embryonic neurons
DNA
cDNA amplification Gel purification of DNA fragments Genomic DNA preparation Phenol extraction from low melt agarose Plasmid DNA prep Silica clean-up of DNA Worm genomic DNA prep In gel-o ligations Cosmid Preparation Cosmid Vectors Codes for C. elegans
RNA
aRNA amplification Joes mRNA preparation Nematode genomic DNA isolation RNA isolation (TRIZO method) RNA preparation RNA synthesis (in vitro transcription) Total RNA preparation Yale RNA preparation
PCR
PCR analysis of dead eggs from C. elegans Polymerase Chain Reaction Single Worm PCR #1 Single Worm PCR #2 Single egg PCR reactions
Proteins
Affinity Purification of Antibodies to GST-fusion Proteins Bacillus thuringiensis (Bt) Toxin Purification Electrotransfer of proteins (electroblotting) Immunoprecipation Preparation of total SDS-soluble nematode proteins Preparation of GST-capping protein Protease inhibitors Purification of MBP Fusion Protein Purification of GST-Fusion Protein SDS- Page
Transforming Worms
Integrating extrachromosanal array into C. elegans chromomosomes Large Scale Transformation by microparticle bombardments
RNAi
96-well format RNAi in liquid culture Bacteria mediated RNAi protocol dsRNA preparation for RNAi by injection in C. elegans (PDF) Making RNAi primers RNAi experiments Double Stranded RNA interference (RNAi ) in C. elegans Microinjection of RNA into C. elegans RNA injection
Staining
Antibody staining
Antibody purification and Testing C. elegans P-granule antibody staining C. elegans Hermaphrodite or Embryo Antibody Staining: Seydoux and Dunn Method DCF staining Freeze- fracture for Immunofluorescence of Embryos Immunofluorescence of C. elegans Embryos Serotonin Antibody Straining – Short version – Long and detailed version Solutions for Anti-Serotonin protocol PAR1-staining Preabsorbing Antisera with C. elegans Acetone Powder
Fixation
Electron Microscopy (EM) Fixation of C. elegans Embryos EM fixation of embryos Fixation Fixation and Permeabilization Fixation and staining parameters Standard Osmium/Glutheraldehyde Fixation Microwave fixes
Bacillus Thuringiensis Crystal Staining C. elegans: Whole Animal DAPI Staining Double dye filling Dye filling to stain amphid and phasmid neurons In situ hybridization Gonad dissections, fixation; DAPI staining, antibody staining, germline in situ hybridization Mms integration Nematode dye filling Phalloidin Staining RNA in situ hybridization of dissected gonads Staining C. elegans for B-galactosidase
Microscopy
Electron Microscopy (EM) Fixation of C. elegans Embryos EM fixation of embryos Fluorescent dissecting stereo microscope Laser Ablation System
Other techniques
Fermentor Gonad dissections Lethal phase determination Mms integration Quick and easy yeast transformation The Electropharyngeogram
