Team:Illinois/Protocols

Protocols

This page describes protocols or includes links to protocols used in our project. Recipes used are also listed in a separate section. Protocols are organized by category. They were collected from Open Wet Ware, our advisors, and labs around the University of Illinois campus.

Standard

• Bacterial Culture Basics (media/plate preparation, growth, streaking, and storage)

• Preparing Cryo stocks

• Transformation by Heat Shock Protocol

• Agarose Gel Electrophoresis Protocol

• Annealing Oligonucleotides Protocol

• PCR Cleanup Protocol (QIAquick PCR Purification Kit) -- Click on "QIAquick Spin Handbook - English (PDF)" under the "Handbooks" header

• Miniprep Protocol (PerfectPrep Spin Mini Kit)

• Miniprep Protocol (PureYield Plasmid Miniprep System)

• BioBrick information

sRNA Characterization

Taken from: A Green Fluorescent Protein (GFP)-Based Plasmid System to Study Post-Transcriptional Control of Gene Expression In Vivo: Johannes H. Urban and Jörg Vogel, 2009

This protocol is used to characterize an E. coli small RNA and its ability to downregulate a reporter gene (GFP). The small RNA is cloned out of the E. coli chromosome and inserted into a high-copy number plasmid (pJU-334). The sRNA's target sequence is also cloned out of the E. coli chromosome and inserted into a low-copy number plasmid (pXG-10). Both plasmids are then transformed into E. coli Top10F' cells. Successful sRNA repression is indicated by a decreased level of fluorescence in the cells.

sRNA characterization procedure

Recipes

LB Broth

LB Plates

EDTA

TAE Buffer

TBE Buffer

Agarose Gels