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0
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1answer
15 views

How do you obtain specific mRNA transcript levels for comparison from a Hi-Seq dataset?

I have HiSeq data from mice exposed to two conditions. I would like to answer the following question: "Is there a significant difference in mRNA transcript levels when comparing condition A to ...
0
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1answer
57 views

How do high-throughput/NGS sequencers calculate quality scores?

I am confused as to how quality scores are actually calculated by DNA sequencers like Illumina. For each base call, some quality predictor value is computed, based on various properties of the ...
2
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0answers
17 views

how to make plasma cells adhere to the bottom of a microplate?

I am isolating single plasma cells by FACS sorting into 384-well plates, with the intent to assay the supernatant and clone H/L chains from positive wells. The efficiency of the PCR is however low, ...
3
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1answer
99 views

Is there a photobleaching-resistant, cell-permeant viability stain in the far red part of the spectrum?

I am looking for a live-cell–impermeable dye for viability. (The cells cannot be permeabilized and fixed in this experiment.) I would prefer with excitation and emission spectra similar to Cy5, but I ...
12
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1answer
914 views

How to reduce edge effects in cell-based high-throughput experiments?

In high-throughput experiments where cells are cultured, treated, stained, and imaged in 384-well microplates, I frequently see significant edge effects. For instance, the following plot shows cell ...