Purified genomic DNA from blood or tissues is analyzed by PCR for single nucleotide polymorphisms (SNPs) relating to one carbon metabolism, (e.g.: PEMT rs12325817, CHDH rs12676 and CHDH rs9001). While we specialize in these SNPs, others can be analyzed by special request. There may be an extra charge for reagent purchase for special request SNPs.
In most cases, TaqMan® SNP Genotyping Assays will be used to detect SNPs and real time PCR carried out on an Eppendorf Mastercycler ep gradient Realplex4 (Eppendorf).
If you need DNA extracted from your sample, see ‘Genomic DNA Extraction’ service for details. If you are extracting your own DNA, we recommend a column-based method such as Qiagen’s QIAamp DNA Blood Mini Kit (Qiagen). Column methods are efficient, automatable, and have less chance of yielding samples with contaminants that could inhibit downstream applications. DNA should be dissolved in 10mM Tris-HCL pH 8.0, 1mM EDTA (TE) (equivalent to Qiagen’s buffer AE). DNA must be of high quality (A260/280 ratio of ~1.8) and free from contaminating reagents such as ethanol or phenol. To ensure the highest possible quality DNA, a column based method is strongly suggested. Quantitate DNA using the Quant-iT™ PicoGreen ® dsDNA Kit (Invitrogen # P11496).
DNA Sample Preparation Requirements
• Send samples in a 96 well plate (8 by 12).
• Amount & concentration*: 50 µl of DNA at a concentration of at least 20-50 ng/µl (1-2.5 µg total DNA) or more is acceptable.
• If necessary, dilute purified genomic DNA in 10 mM Tris-HCL pH 8.0, 1 mM EDTA (TE).
• Prepare the sample sheet (we will provide this) with the following information: PLATE Name, POSITION, Sample Id, A260, A280,
• 260/280 ratio, DNA concentration (ng/µL), DNA volume in plate (µL), and Total DNA amount (ng).
• One sample sheet for all the plates. Comma separated CSV, Excel or tab delimited txt format is acceptable.
• Samples must be coded to ensure anonymity.
* We will use NanoDrop ND-1000 Spectrophotometer to double check the DNA concentration.
Standard Operating Procedures
A copy of the IRB under which the samples were collected is required and methods must include processing and genotyping of samples. Samples not following the above requirements will not be processed.
Have Questions? Contact Us.
Saroja Voruganti, PhD
Core Director, Nutrigenetics Core
T: (704) 250-5009