The QB3 MacroLab is a core service facility that operates on a recharge basis with varying rates for UC users and those not affiliated with UC.
Automated methodologies substantially decrease the price per sample, however not everyone needs a plate of 96 clones or proteins, and we are happy to take on any size project. Many projects begin with creating expression clones and culminate in the delivery of purified proteins. The MacroLab also can assist in any number of steps in this process.
Researchers requesting MacroLab services vary widely. Biochemistry and structure labs often request our 96-well services to screen for well-behaved proteins amenable to purification. Genetic labs have requested various types of 96-well gene libraries. Labs that do not specialize in our methodologies, such as computational and drug screening labs, have requested MacroLab to perform all of their cloning and protein purification. In addition, many of our in-house labs save substantially on our high-quality reagents.
High-throughput cloning in the MacroLab is carried out using LIC (Ligation Independent Cloning) because of its high efficiency. Target open reading frames can be amplified from genomic, cDNA, or plasmid DNA. We have a large variety of bacterial and baculovirus expression vectors on hand to accommodate most needs. We have, in many cases, modified a customer’s favorite plasmid to be LIC compatible in order to perform 96-well cloning.
Quick-change mutagenesis is carried out in high-throughput format. Sets of 24 or 96 quick-change reactions are performed in 96-well plates using a single-oligo quick-change methodology.
LIC cloning and quick-change mutagenesis may also be requested for very small sample numbers. Often, projects involving fewer samples are highly customized to meet the user’s needs. Methodologies including traditional restriction enzyme digestion and ligation, gene synthesis, or SLIC (Sequence and Ligation Independent Cloning) are routinely employed to construct the plasmids requested.
Protein expression in E. coli is routinely performed in 96-well format using 1mL cultures. We have optimized parameters for several temperatures using both IPTG and auto-induction. Larger 1L flask cultures and 4L fermenters are also available for larger preps.
High-throughput nickel and streptavidin purifications are available for cultures grown in a 96-well plate. Multiple liter protein purification involving affinity, ion exchange, and size exclusion chromatography is performed routinely.
Reagents including Cas9, competent cells, Phusion polymerase, mastermixes, and TEV protease are available.
See the attached pdf for a detailed description of MacroLab’s services. As described above, most projects we perform vary substantially in cost due to the sample number and the techniques required. Please contact us to discuss any project you have in mind, we are happy to assist in your project’s design, and will provide you with a quote.
The MacroLab welcomes input and suggestions of methodologies and reagents that would enhance the facility and opportunities within QB3. Please contact Chris Jeans with questions, suggestions, and comments.