Sample Preparation Guidelines
We accept samples from all over the world, but it is important to follow the guidelines below:
· The minimum number of samples for DNA extraction is 94.
· For young leaf tissues: Collect young leaf samples without necrotic lesions, preferably 21-30 days old. Cut 4 inches from the tip of the leaves and fold only twice. Place inside glassine bags. Labeling of bags containing the leaf tissue should follow this format: date/ project or study name/ bag number (if several)/ sample owner.
ie. Requestor: Juan Dela Cruz: 23May2013/GrainQ/1/JDC
· For lyophilized tissues: Place 4 inches of lyophilized leaves and fold only twice in a 2ml deep well plates. Labeling of plates containing the lyophilized tissues should follow this format: date prepared/ project or study name/ plate number (if several)/ sample owner.
ie. Requestor: Juan Dela Cruz: 23May2013/Grainq/Plate1/JDC
Note: For DNA extraction, we only accept samples which will also be processed for SNP Genotyping.
· For DNA extraction of your leaf samples, for best results we recommend the use of Modified CTAB Method with RNAse.
· For optimum DNA yield, collect leaf tissues from 3-week to one-month old plants.
· Check quality of DNA samples on 0.8-1% agarose gel. Bands should be clear. Lanes with white smear indicates presence of RNA in the samples.
DNA quality check on 1% agarose gel.
· Quantify DNA using NanoDrop Reader or other spectrophotometer.
· Pure DNA should have 260/280 nm ratio of approximately 1.8-2.0 and a 260/230 ratio of ˃1.5.
· Standardize DNA concentration to 50ng/µl by diluting it with sterile NanoPure water or TE buffer.
· Dispense at least 20 µl of 50ng/µl DNA in a 96-well plate. The use of sticky seals is not sufficient to seal the plate; we recommend the use of a thermal sealer. The use of Cap Mat sealing is a suitable alternative if heat sealing is not available.
· Don’t forget to attach the picture of the gel and the quantification of the DNA when you submit the SNP Genotyping form.
In the plate dispense samples column-wise and follow this format: date prepared, project/study name, plate number (if several) and initials of sample owner.
Note: Do not forget to fill the Plate Layout in the SNP Genotyping form.
ie. Requestor: Juan Dela Cruz
Plate layout and labeling of DNA samples for SNP Genotyping.
A. SNP genotyping by Illumina BeadXpress:
·The minimum number of DNA samples that can be submitted is 24 per OPA (test)
·For submissions greater than 24 DNA samples, we require multiples of 24 in a 96 well plate.
B. SNP genotyping by Fluidigm 24 SNPs set:
·The minimum number of DNA samples that can be submitted is 188 per assay.
·DNA samples to be submitted should be equal to or multiples of 188.
·The DNA samples should be submitted in 96-well plates.
C. SNP genotyping by Fluidigm 96 SNPs set:
· The minimum number of DNA samples that can be submitted is 94 per assay.
· DNA samples to be submitted should be equal to or multiples of 94.
· The DNA samples should be submitted in 96-well plates