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...performed at Addgene on the A549 cells described in our sample data , each 96-well plate gave rise to 15–20 monoclonal...solution will be used to seed the 96-well plate. Sample Calculation First, use a hemocytometer to quantitate...plate for individual cells, which should be on the same focal plane. Scan the entire plate and make note...highest or lowest transgene expression ( Figure 2 ). Sample Data Figure 1: Generation of monoclonal cell lines...

...gel extraction of either the original sample or the amplified sample, followed by reamplification of the...tens to millions of different plasmids in a single sample. They are often used for screening, barcoding, ...appear to contain sufficient DNA for use, pool samples, and continue with analysis. “Sufficient” DNA will...

...before you place the sample at -80°C. Frozen tubes are hard to write on and samples stored for long periods...glycerol. You can prepare the glycerol stock the same time you prepare your plasmid DNA. In the morning...

...Pipetting Technique Learn how to pipette multiple samples at once using a multichannel pipette and avoid ...bands out of the agarose gel and purify the DNA samples. This is a commonly used technique for molecular... Your guide to the MTA process when requesting samples from Addgene Technology Transfer How to Order A...

...optimal fixation method will vary depending on the sample type and the protein of interest. You may need ...expected and specific, include a positive control sample that you know expresses the protein, such as cells...the protein of interest, and a negative control sample such as cells that do not express the protein of...

...of DNA Samples Add an equal volume of TE-saturated phenol-chloroform to the aqueous DNA sample. Pro-Tip...remaining contaminant proteins and RNase A from the DNA sample. When phenol is mixed with the aqueous solution...

...absorbed. The spreading of cells can be done in the same way as the antibiotic, using either a bent micropipette...over-agar for selection of transformed E. coli . Sample Data Selection of E.coli on LB-agar using different...

...two different shRNAs that target the same gene can produce the same phenotype will alleviate concerns about...boiling water. If using a PCR machine, incubate the sample at 70°C for 10 minutes then slowly cool to room...

...double digest (digesting with two enzymes at the same time), you will need to determine the best buffer...are digesting a large number of plasmids with the same enzyme(s) (for instance, in a diagnostic digest)...

...protocol outlines the seeding of the cells at the same time as the virus-mediated transduction. For some... titer, take the average of multiple dilutions. Sample Data Figure 1: A549 cells were transduced with ...

...complete + 10 µg/mL polybrene. Note, this is just a sample of possible dilutions. You may want to try higher...monoclonal lines from the early polyclonal populations. Sample Data Figure 1: Generation of monoclonal cell lines...

...primer is used to amplify a eukaryotic genomic DNA sample. However, a primer should not be too long (> 30...

...just after base 70 there are multiple peaks in the same location. Looking at the trace file will give you...

...have different leader sequences, but operate on the same principle (homologous to the first G on 3'-5' strand...

...centrifuge to separate different components in a liquid sample Using a Light Microscope Learn about the parts ...

... titer, take the average of multiple dilutions. Sample Data Figure 1: 293T cells were transduced with ...

...24–48 h before checking for protein expression. Sample Data Legend: Lenti-X 293T cells were transfected...

... °C as soon as possible to avoid loss of titer. Sample Data Figure 1: 293T cells were transfected with...

... volumes you need to pipet. If you are using the same template DNA for all your reactions, the Template...