Common Procedures

Use the table of contents to navigate this repo of common lab procedures. These protocols were adapted 2020-2021 by Chase Weidmann.

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Table of Contents

• Procedure Template
• General
• NGS
• Gateway Cloning
• Lentivirus Stables
• Chemical Probing
• Mutational Profiling
• Proximity Labeling
• Mass Spec Prep

General

• BCA Assay
• DNA Agarose Gel
• Reseal Inverse PCRs
• Making frozen cell stocks
• Gibson Assembly
• Restriction Digest
• SDS-PAGE for protein
• Standard PCR
• DNase treatment of RNA
• Trizol RNA Purification
• Western Blotting

NGS

• Second Strand Synthesis
• G-50 Buffer Exchange
• SPRI Bead Cleanup and Size Selection
• Nextera XT Library Prep
• Nextera Plasmid Library Prep
• 2-step Illumina Library Prep

Gateway Cloning

• pENTR Creation
• Multisite Assembly

Lentivirus Stables

• Virus Production
• Infection and Selection

Chemical Probing

• SDA Crosslinking
• OOPS RNP Enrichment
• In-cell SHAPE probing
• Cell-free DMS probing
• In-cell DMS probing
• Cell-free DMS probing

Mutational Profiling

• MaP RT with SuperScript II
• MaP RT with Marathon

Proximity Labeling

• miniTURBO Biotinylation
• RIPA Whole Cell Lysis
• miniTURBO Streptavidin Pulldown
• In-cell DSP Crosslinking
• HaloTag Mammalian Cell Lysis
• HaloTag Pulldown

Mass Spec Prep

• On-bead Digestion
• Bead Reduction and Elution
• C18 Column Cleanup
• Ethyl Acetate Cleanup
• Peptide Quantification