DNA/RNA Sample Preparation and Quality Control
It’s very important that users provide us DNA or RNA samples at required quality and quantity. Please refer to the Services section for specific requirements of sample submission. Here are some general guidelines for sample preparation and quality control:
Total RNA preparation
- Total RNA samples for sequencing or microarray analysis should be free of proteins, DNA, phenol, ethanol, and salt.
- We recommend columns (such as Qiagen RNeasy columns) for RNA preparation
- If TRIzol is used to extract RNA from tissues, we recommend a cleanup procedure with columns
- DNase treatment is recommended
We recommend the NanoDrop spectrophotomer for quick quantitation. NanoDrop at our center is available free of charge.
RNA quality assessment
The quality of total RNA samples is very important in RNA-Seq and microarray experiments. When you provide us total RNA samples, we always perform RNA integrity check on Agilent 2100 Bioanalyzer. RNA integrity number (RIN) at 8.0 or higher is required by most protocols.
Genomic DNA Preparation for SNP genotyping and sequencing
- Genomic DNA samples should be free of PCR inhibitors (such as heme and high concentrations of EDTA and salt)
- DNA samples must not be contaminated with other sources of DNA. This is particularly important in SNP microarrays, as a universal amplification step is involved. Any contaminant DNA will also be amplified and affect the data.
- For Affymetrix SNP arrays, DNA samples should be in reduced EDTA TE buffer (10 mM Tris, 0.1 mM EDTA, pH 8.0)
- QIAamp Kit from Qiagen has been used successfully for DNA preparation
- NanoDrop Spectrophotometer
- Qubit Florometer
Genomic DNA Quality Assessment:
- DNA samples must not be highly degraded. We recommend quality check on 1% agarose gel. High quality genomic DNA should give a major band at 10-20 kb on the gel.
- We also recommend that users perform a regular PCR amplification on the genomic DNA samples with a pair of primers of users’ choice. This is to ensure that genomic DNA samples are free of PCR inhibitors.