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question about normalization of Nimblegen expresssion array

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xiao wu ▴ 10

@xiao-wu-3398

Last seen 10.3 years ago

Hi all, I am using the Nimblegen expression microarray. Each array has 4 different bacteria genome, and the position of probes are randomly assigned. I have two groups: control and treatment, and I used the mixed RNA from 4 bacteria under each condition to apply to array. However, the quantity of RNA from 4 bacteria may be different, and the difference is not consistent between control group and treatment group. Should I normalize separately for each bacteria? And if yes, what's the proper way to do that? Can I take the raw calls file provided by Nimblegen for each bacteria and then apply quantile normalization? Thanks for help. Xiao [[alternative HTML version deleted]]

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ADD COMMENT • link written 15.7 years ago by xiao wu ▴ 10

Gordon Smyth · Answer 1 · 2016-10-28

Bacteria expression are the most widely used protein expression systems for their rapid growth, high yield, ease of manipulation and scale-up. The most widely used bacteria host is of course Escherichia coli. E. coli is the most established tool and is almost always the first choice if applicable.

Another bacterium, Bacillus subtilis, is an increasingly popular protein expression host. It is a FDA designated Generally Regarded As Safe (GRAS) organism. B.subtilis is a Gram-positive bacterium, which makes this system free of endotoxin concern. It also has much less codon bias than E. coli. Moreover, it secretes protein directly into the culture media, which highly simplifies subsequent protein purification steps. All the above features make it an attractive system for the production of soluble and properly folded recombinant proteins at both laboratory and industrial levels.