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        丁香实验推荐阅读
        Joe mRNA prep

        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 ...

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        Yale RNA prep

        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 ...

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        Immunofluorescence

        Immunofluorescence of C. elegans EmbryosMaterials:fixative dependent on antigen (common: 4% formaldehyde or 100% MeOH) post-fixative dependent on antigen (usually MeOH) PBS 1X PBS PBST PBS + detergent ...

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        DAPI Staining

        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 ...

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        Double dye filling (using DiO and di-4 ANEPPS)

        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 ...

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        Dye Filling to Stain Amphid and Phasmid Neurons

        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 ...

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        Fixation and Staining Parameters

        Fixation and Staining ParametersAntigen Fixation Fixation time AntibodyDilutionRefmicrotubules 100% MeOH 5 min @ RT DM1a1:500actin 4% Formaldehydundefined 8 min @ RT C41:5001POD-1 Formaldehyde or MeOundefined 8 min ...

        丁香实验推荐阅读
        EM Fixation of Embryos

        EM Fixation of EmbryosAdapted following advice from Andy Fire and Jim Priess1. Mount embryos on slides as if for DIC except agar has fixative. Agarose Fixative 3ml 2% Agarose : 1.5% LGT Agarose + .05% ...

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        Drosophila Media Recipes and Methods

        Bloomington No-Plug Food Prep Method Standard medium in use at Bloomington Hard agar medium in use at Bloomington Cornmeal Sucrose Dextrose Yeast and 2-Acid Medium Cornmeal-molasses-yeast medium Co ...

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        Gonad Dissections

        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 ...

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        Gonad Fixation

        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 ...

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        In situ hybridization

        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 ...

        丁香实验推荐阅读
        FREEZE-CRACK b-GALACTOSIDASE STAINING PROTOCOL

        FREEZE-CRACK b-GALACTOSIDASE STAINING PROTOCOLFIRE LAB1. Prepare humidity chamber: line large square plate with humid paper towels and place 4 5ml plastic pipets across chamber to support slides.Place ...

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        Preparation of single-stranded probes from cloned cDNA

        Preparation of single-stranded probes from cloned cDNA(from Patel and Goodman 1992)a) 2-5 µg of plasmid DNA containing the cDNA insert is linearized using an appropriate restriction enzyme. For antise ...

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        MMS integration

        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 ...

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        Nematode dye filling

        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 ...

        丁香实验推荐阅读
        Whole-mount in situ hybridization for the detection of RNA in C. elegans embryos

        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 ...

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        CONVENTIONAL TWO STEP FIXATION [See D.H. Hall, 1995]

        "CONVENTIONAL" TWO STEP FIXATION 1995 1. Wash animals in M9 and anaesthetized in 8% alcohol in M9 5’ 2. 2.5% glutaraldehyde 1% paraformaldehyde in 0.1M sucrose 0.05M cacodylate on ice 2 hours 3. 3 ri ...

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        Phalloidin Staining worms

        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 ...

        丁香实验推荐阅读
        Staining C. elegans for ß-galactosidase

        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 ...

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