RNAi In Situ

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View the entire protocol, "In Situ Hybridization of Whole-Mount Mouse Embryos with RNA Probes: Hybridization, Washes, and Histochemistry," at BioSupplynet.com

Thomas Lufkin This protocol was adapted from "Techniques for Visualizing Gene Products, Cells, Tissues, and Organ Systems," Chapter 16, in Manipulating the Mouse Embryo, 3rd edition, by Andras Nagy, Marina Gertsenstein, Kristina Vintersten, and Richard Behringer. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA, 2003. ABSTRACT This is a protocol describing how to perform in situ hybridization on whole mouse embryos. Here we describe the hybridization procedure, and the localization of the DIG-labeled RNA using a conjugate of anti-DIG Fab antibody and calf intestinal alkaline phosphatase. Enzyme activity of the reporter is detected by a color reaction, resulting in the formation of a water-insoluble purple/blue precipitate.

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Products Available for this Protocol

Protocol Material Description	Product #	Product Name	Add to Cart
Tris-Cl	T5941	Trizma® hydrochloride, for molecular biology, Biotechnology Performance Certified, cell culture tested, ≥99.0% (titration)
NaCl	S3014	Sodium chloride, for molecular biology, ≥98% (titration)
EDTA	E5134	Ethylenediaminetetraacetic acid disodium salt dihydrate, for molecular biology, ≥99%
PBS	P5368	Phosphate buffered saline, pH 7.4, Biotechnology Performance Certified
Proteinase K	P6556	Proteinase K from Tritirachium album, lyophilized powder, ≥30 units/mg protein
MgCl2	M1028	Magnesium chloride solution, for molecular biology, 1.00 M±0.01 M
Tween 20	P9416	TWEEN® 20, for molecular biology, viscous liquid
Methanol	34860	Methanol, CHROMASOLV®, for HPLC, ≥99.9%
H2O	W4502	Water, Molecular Biology Reagent
Blocking Reagents
SSC	S8015	SSC Buffer
Formamide	F7508	Formamide, for molecular biology, ≥99.5% (GC), liquid
CHAPS	C5070	CHAPS, SigmaUltra, ≥98% (TLC)
Heparin	H4784	Heparin sodium salt, ~170 USP units/mg
Maleic acid	M0375	Maleic acid, ReagentPlus®, ≥99%
Paraformaldehyde	P6148	Paraformaldehyde, reagent grade, crystalline
KCl	P9541	Potassium chloride, for molecular biology, ≥99%
Na2HPO4	S3264	Sodium phosphate dibasic, for molecular biology, ≥98.5% (titration)
KH2PO4	P9791	Potassium phosphate monobasic, for molecular biology, ≥98%
CaCl2 • 2H2O	C7902	Calcium chloride dihydrate, cell culture tested, insect cell culture tested, ≥99%
MgCl2 • 6H2O	M2393	Magnesium chloride hexahydrate, cell culture tested, insect cell culture tested
BM purple AP substrate	B3679	BCIP®/NBT-Purple Liquid Substrate System for Membranes
Sheep serum	S2263	Sheep Serum, USA origin, sterile-filtered, cell culture tested
Sodium azide	S8032	Sodium azide, SigmaUltra

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