vitro Protocols

Category: RNA Protocols: In Vitro Transcription T7 SP6 T3 Protocols

Ribomax T7 In Vitro Transcription Protocol. Synthesis of mg quantities of RNA from a T7 promoter. Promega. - [Read Ribomax T7 In Vitro Transcription Protocol]

Category: RNA Protocols: In Vitro Transcription T7 SP6 T3 Protocols

Riboprobe® in vitro Transcription Systems Promega. For the in vitro synthesis of RNA. For large quantities of RNA, use Ribomax very good kit. - [Read Riboprobe in vitro Transcription Protocol]

Category: Cell Biology and Cell Culture: In Vitro Reconstitution Protocols

In vitro transcription systems includes instructions for use of products P1420, P1430, P1440, P1450 and P1460.Includes information and protocols on RNA Transcription in vitro. Information on DNA Template Preparation;Synthesis of High-Specific-Activity Radio labeled RNA Probes;Determining Percent Incorporation and Probe Specific Activity;Removal of the DNA Template Following Transcription;Removal of unincorporated nucleotides;Synthesis of large amounts of RNA;Capping RNA for in vitro translation. - [Read Riboprobe In Vitro Transcription Systems]

Category: Cytogenetics Protocols: DIG Biotin Labeled Probe Protocols

Protocol for RNA labeling by in vitro transcription of DNA with DIG, Biotin or Fluorescein RNA Labeling Mix. - [Read RNA Labeling by In Vitro Transcription of DNA with DIG, Biotin or Fluorescein RNA Labeling Mix]

Category: Cytogenetics Protocols: RNA In Situ Hybridization Protocols

Protocol for RNA labeling by in vitro transcription of DNA with DIG, Biotin or Fluorescein RNA Labeling Mix. A PCR fragment that has the appropriate promoter ligated to its 5’-ends can also serve as a transcription template. The procedure described incorporates one modified nucleotide (DIG-, Biotin-, or Fluorescein-UTP) at approximately every 20 – 25th position in the transcripts. - [Read RNA Labeling by In Vitro Transcription of DNA with DIG, Biotin or Fluorescein RNA Labeling Mix]

Category: Nucleic Acid Modification Protocols: DIG Biotin Labeled Probes Protocols

Protocol for RNA labeling by in vitro transcription of DNA with DIG, biotin or fluorescein RNA labeling mix. - [Read RNA Labeling by In Vitro Transcription of DNA with DIG, Biotin or Fluorescein RNA Labeling Mix]

Category: Cell Biology and Cell Culture: In Vitro Reconstitution Protocols

Information for protocol using single-tube, coupled transcription/translation reactions for eukaryotic in vitro translation. Includes information on: Translation Procedure; Positive Control Translation Reactions Using Luciferase; Cotranslational Processing Using Canine Pancreatic Microsomal Membranes; Post-Translational Analysis; Positive Control Luciferase Assays; Composition of Buffers and Solutions; Luciferase SP6/T7 Control DNAs - [Read Single-tube Coupled Transcription/Translation Reactions for Eukaryotic In Vitro]

Category: Pharmacology and Toxicology Protocols: Drug Discovery

In Vitro ABC Transporter Assays for Hepato-Biliary DMPK; ABC transporter assays; Uptake transporter assays; Transporter assays for enteral absorption PK; Transporter Assays for Blood-Brain-Barrier PK; In Vitro Transporter Assays for renal excretion. - [Read SOLVO In Vitro Transporter Assay Technology]

Category: Reporter Gene Assays: Luciferase Assay Protocols

Protocol describes a split luciferase complementation assay used to study the interaction of proteins in cells. In the split protein strategy, a single reporter protein/enzyme (firefly luciferase [Fluc]) is cleaved into amino-terminal and carboxy-terminal halves; each half is fused to one of two interacting proteins, X & Y. Physical interactions between the two proteins reconstitute the functional reporter protein, leading to enzymatic activities that can be measured by in vitro or in vivo assay - [Read Split Luciferase Complementation Assay for Studying Interaction of Proteins X and Y in Cells]

Category: Cell Biology and Cell Culture: In Vitro Reconstitution Protocols

Small ubiquitin-like modifier (SUMO) is a small molecule, but has a variety of regulatory functions in cells. SUMO modification is involved in transcriptional regulation, subcellular localization, and protein-protein interactions. SUMO conjugation requires sequential E1-dependent activation, E2-dependent conjugation, and E3-dependent ligation steps. Protocol includes: In vivo and in vitro SUMOylation assay and deSUMOylation assay. - [Read Sumoylation and Desumoylation Assays for a Chromatin-Remodelling Complex In Vivo and In Vitro]

Category: Nucleic Acid Purification Protocols: RNA Isolation Protocols

Protocol for total RNA isolation. This method is fast and works well for preparation of total RNA.  We typically use total RNA for quantitative S1 nuclease analysis of endogenous (or in vitro) gene expression, and this method works well for this purpose. - [Read Total RNA isolation (Guanidinium Thiocyanate-Phenol-Chloroform Extraction) Protocol]

Category: Microarray Protocols: Microarray Hybridization Protocols

Protocol details the preparation of fluorescently labeled target samples and hybridization of these samples to a microarray of Agilent inkjet-deposited cDNAs.  The procedure requires a minimum of 5 mg of purified total RNA as starting material. Includes: First Strand cDNA Synthesis; Second Strand cDNA Synthesis; cDNA Cleanup and Precipitation; In vitro Transcription; Cleanup and Quantification of in vitro Transcribed RNA; Fluorescent Labeling of the Target Samples. - [Read Transcript Profiling by Microarray Analysis Protocol.]

Category: Transfection Protocols: Calcium Phosphate Transfection Protocols

This protocol describes two transfection methods for expressing GFP-tagged actin in primary neurons. The lipid reagent DOTAP (Roche Diagnostics) method produces actin-GFP-expressing hippocampal neurons that survive well during long periods in culture. The calcium phosphate method can be used to transfect neurons that have already been growing on coverslips in vitro. Transfected cells suitable for imaging can be obtained in cultures up to 15 days in vitro. - [Read Transfecting Cultured Hippocampal Neurons with an Actin-GFP Plasmid]

Category: Mouse Protocols: Mouse Reproductive System Surgery Protocols

Protocol describes a procedure for uterine transfer, which is used for chimera production. The method is based on extensive work which resulted in the first successful development and birth of in-vitro-cultured mouse embryos. It is best to practice this procedure first on a cadaver and then on an anesthetized 2.5-days post coitum (dpc) pseudopregnant mouse using blue Affigel beads rather than embryos. - [Read Uterine Transfer Protocol]

Category: Xenopus Protocols: Xenopus Egg Extracts Protocols

Protocol describes the preparation of Cytostatic Factor (CSF) from frog eggs. CSF can be used in in vitro spindle assembly assays. Includes: Frog injections; Preparation Before Starting the Extract Preparation; Extract Preparation. - [Read Xenopus Egg CSF Extract Preparation Protocol]