The GSL can provide the following services in high-throughput to support your research needs.
Blood Fractionation from fresh blood (for plasma/buffy coats or serum/blood clots)
Blood samples are fractionated by centrifugation. Samples provided in heparin (green-top), EDTA (lavendar-top), or ACD (yellow-top) vacutainers are separated into plasma , buffy coat layer, and red blood cell layer. Samples provided in red tops (either with or without coagulants) are separated into serum and blood clot layers. Plasma or serum is aliquoted into microfuge tubes. Buffy coats are collected and subsequently extracted for DNA (additional charge applies for DNA extraction).
DNA Extraction from 10ml Whole Blood or ~3ml Buffy Coat
DNA is extracted from either buffy coat or whole blood using the Gentra Puregene Blood Kit. Through a series of centrifugations and additions of lysis reagents, the cells are lysed and pelleted. RNase A Solution is added for RNA-free DNA. Protein contaminants are removed by salt precipitation. Genomic DNA is then precipitated with alcohol and dissolved in hydration solution (1 mM EDTA, 10 mM Tris·Cl pH 7.5).
DNA Extraction from Saliva
DNA is extracted from saliva collected by Oragene saliva collection kits. Through a series of centrifugations and addition of Oragene’s purifier, the cells are lysed and the DNA is purified. The DNA is then ethanol precipitated and dissolved in hydration solution (1 mM EDTA, 10 mM Tris·Cl pH 8).
DNA Extraction from other tissue types
We also extract DNA from other tissue types including isolated cells and tissues upon request and after review. DNA extraction from isolated cells are done using the Gentra Puregene Kit. DNA extraction from tissues, glands (e.g. Labial glands) are done using the Qiagen AllPrep DNA/RNA Mini Kit. Please contact us in advance for pricing if you are interested in DNA extractions from tissues.
Whole Blood PAXgene DNA Service
The PAXgene Blood DNA Kit allows isolation of DNA from 8.5 ml of human whole blood collected in PAXgene Blood DNA Tubes. Cell lysis buffer is added to the blood to lyse red and white blood cells. Cell nuclei and mitochondria are pelleted by centrifugation, washed, and resuspended in digestion buffer. Protein contaminants are subsequently removed by incubation with protease enzyme. DNA is precipitated in isopropanol, washed in 70% ethanol, dried, and resuspended in resuspension buffer.
Whole Blood PAXgene RNA Service
The PAXgene Blood RNA Kit in conjunction with the PAXgene Blood RNA Tubes allows for the collection of blood and simultaneous stabilization of cellular RNA. The PAXgene Blood RNA Kit allows for purification of total RNA from 2.5 ml of human whole blood collected in a PAXgene Blood RNA Tube. Nucleic acids are first pelleted by centrifugation and subsequently washed and resuspended. Protein digestion is carried out through an incubation with buffers and proteinase K. The cell lysate is homogenized and residual cell debris is removed through centrifugation through the PAXgene Shredder spin column. Ethanol is added to the supernatant of the flow-through fraction and the lysate is applied to a PAXgene RNA spin Column and undergoes another centrfigutaion step, selectively binding the RNA to the PAXgene silica membrane. Through a series of washes and centrifugation, the contaminants are removed. DNase I is also added to remove trace amounts of bound DNA. After the series of washes, RNA is eluted in elution buffer and heat-denatured.
Additional Services for Biospecimen Processing
The GSL can also provide high-throughput DNA/RNA quantitation, normalization, plating, and both transfer (to outside laboratories) or storage of samples. Quantitation is done using the Nanodrop ND-1000 or using the Tecan Infinite 200 Plate reader. We also offer computer tracking of stored samples and investigator-access through a web-based lab database through our Biospecimen Processing Service.
The rates for these services can be found here.
If you have any questions regarding our Biospecimen Processing Service, please contact:
Hong Quach (Illumina Arrays and Biospecimen Processing)
431 Stanley Hall
Berkeley, CA 94720-3220