How Should I Prepare My Samples?
For all samples, we request that you submit a copy of your quote. This will ensure easier tracking of your project, and will help us minimize the the turnaround time.
DNA Samples
For DNA samples, follow your favorite extraction method, as long as it preserves high- molecular-weight fragments. There should essentially be no migration on a 1% agarose gel run for ~1 hour @ 120V. The best DNA cleanup method is phenol/chloroform extraction followed by ethanol precipitation with several washes (And No, your column is not just as good!). RNAse-treated samples are preferred. Clean samples should have a 260/280 ratio of 1.7 to 1.9, and a 260/230 ratio >2. Mail samples resuspend in TE buffer with a cold pack, or dry the sample down completely and send at room temperature. For genomic DNA projects, submit a minimum of 5 micrograms of DNA at a minimum concentration of 100 nanograms / microliter.
ChIP-seq Samples
For ChIP-seq samples, submit any total amount, provided there is a minimum of 20 nanograms in the 150-250bp size range. Analyze this concentration-by-fragment-length distribution using a BioRad Experion or Agilent Bioanalyzer 2100.
RNA Samples
For RNA samples, we recommend the mirVana extraction kit from Ambion (http://www.ambion.com/catalog/CatNum.php?1560). If you read the protocol, you will notice there are two paths through the protocol, one for total RNA and one for microRNAs. We recommend this kit for both–just follow the correct path for your sample.
Please submit your RNA at a minimum concentration of 500 nanograms / microliter. The 260/280 ratio of your RNA sample should be >1.9, and your sample should be resuspended in nuclease-free or DEPC water. RNA samples should be mailed overnight on dry ice, or pelleted and dried to be sent at room temperature. Pelleting is the preferred method for international samples.
For microRNA projects, submit at least 20 micrograms of total RNA. For whole-transcriptome projects, submit at least 5 micrograms of total RNA (10 micrograms is better, however we know this might not be possible for all projects). Cofactor will perform the microRNA or PolyA selection in-house, and even with the best methods, recovery is quite low. Thus, it is prudent to submit as much RNA as possible. Cofactor has been able to create many Ribo-depleted SOLiD whole-transcriptome libraries from as little as 1 microgram total RNA, such as that from of Laser Capture Micro-dissection (LCM). However, the recommended 10 microgram input will likely yield even more quantitative libraries and ensures less risk of library failure.
On a gel, high-quality RNA should have prominent 18S and 28S bands and a prominent smear from .5 to 12kb. On an Agilent Bioanalyzer 2100, RNA should have an RNA Integrity Number (RIN) >=8.
