KS USDA Sample Collection and Submission Protocol For Seeds:
1. Always contact Guihua or Paul BEFORE collecting samples so that the
appropriate sample amounts and methods are used.
2. Always contact our lab BEFORE sending samples so that we can make sure
they arrive safely and are handled properly and promptly.
3. Send 5 or more seeds per line.
4. Label each seed packet clearly and place in them in sample order.
5. Since we must plant the seeds, grow the seedlings, and harvest the tissue prior
to DNA extraction, please expect a delay of 1 month longer than usual before
data is returned.
6. We always work with 96-well sample plates. If you have fewer than 96
samples, you should replicate them to make better use of the sample plate.
7. Always leave the 96th sample empty. It will be the last well (H12) in a 96 well
plate so we can include 1 internal control sample.
8. Always email a sample list in an excel file including: sample plate name and
number, well positions for each sample, sample name, traits of interest,
markers to use for each trait. Other info is helpful, such as sample generation,
pedigree, expected genotype for each marker (positive, negative, segregating,
etc.). See the Sample Excel file.
9. Always include samples of parent lines, known positives, or other controls.
10. Replication of parents or controls is helpful.
11. If you will be requesting newly published markers or non-published markers,
send the protocols 1 or 2 months prior to the samples.
12. Always include a job name.
KS USDA Sample Collection and Submission Protocol For Half-Seeds:
1. Always contact Guihua or Paul BEFORE collecting samples so that the
appropriate sample amounts and methods are used.
2. Always contact our lab BEFORE sending samples so that we can make sure
they arrive safely and are handled properly and promptly.
3. Send 1 to 5 half-seeds per well. 1 half-seed per well is sufficient for testing
parents with up to 40 markers. Multiple half-seeds per well would be useful
for testing lines while reducing sample numbers. Obviously, send only the
endosperm end of the seed and keep the embryo end for planting.
4. Collect seeds into the approved 1 ml plates with mats.
5. There is no need for dry ice while shipping. Samples can be stored at room
temperature for short periods (~14 days) before sending them.
6. We always work with 96-well sample plates. If you have fewer than 96
samples, you should replicate them to make better use of the sample plate.
7. Always leave the last well (H12) EMPTY so we can include 1 internal control
sample.
8. Always email a sample list in an excel file including: sample plate name and
number, well positions for each sample, sample name, traits of interest,
markers to use for each trait. Other info is helpful, such as sample generation,
pedigree, expected genotype for each marker (positive, negative, segregating,
etc.). See the Sample Excel file.
9. Always include samples of parent lines, known positives, or other controls.
10. Replication of parents or controls is helpful.
11. If you will be requesting newly published markers or non-published markers,
send the protocols 1 or 2 months prior to the samples.
12. Always include a job name.
For half seeds, use only 1ml 96-Deepwell plates from bioexpress.com or VWR:
From Bioexpress (www.bioexpress.com)
1 ml Deep-Well Riplate, Polypropylene, Non-Sterile, 20/case, #P-8700-1. (NOTE:
Use Beckman mats. Does NOT work with VWR mats.)
From Beckman (www.beckmancoulter.com)
Cap Mats for 1ml 96 well plate, Nonsterile, Case of 12, #267002 (NOTE: Use these
for the plates from Bioexpress, DO NOT USE BECKMAN PLATES, THEY
LEAK!)
From VWR (www.vwrsp.com)
1ml Polypropylene Deep Well Microplates, Nonsterile, Case of 50, #40002-009
Mat Lids For 1.0 mL Microplates, Nonsterile, Case of 50, #40002-001
KS USDA Sample Collection and Submission Protocol For Fresh Tissue On
Dry Ice:
1. Always contact Guihua or Paul BEFORE collecting samples so that the
appropriate sample amounts and methods are used.
2. Always contact our lab BEFORE sending samples so that we can make sure
they arrive safely and are handled properly and promptly.
3. Always collect samples into approved 96-well plates.
4. We always work with 96-well sample plates. If you have fewer than 96
samples, you should replicate them to make better use of the sample plate.
5. Always leave the last well (H12) EMPTY so we can include 1 internal control
sample.
6. Always email a sample list in an excel file including: sample plate name and
number, well positions for each sample, sample name, traits of interest,
markers to use for each trait. Other info is helpful, such as sample generation,
pedigree, expected genotype for each marker (positive, negative, segregating,
etc.). See the Sample Excel file.
7. Always include samples of parent lines, known positives, or other controls.
8. Replication of parents or controls is helpful.
9. If you will be requesting newly published markers or non-published markers,
send the protocols 1 or 2 months prior to the samples.
10. Always send a UNIFORM amount of sample across wells.
11. Always label plates clearly with the plate name or number and the job name.
12. Keep the samples cold during collection (keep the plate on ice).
13. Collect 4 to 6 leaves (50 to 200mg total) of 2 cm length each (longer pieces do
not grind as well). Use only very young tissue, the 3 leaf stage works well. Do
not put more than 200mg of fresh tissue into a well.
14. Use dry ice for shipping. Samples should be collected and shipped on the
same day, or should be frozen right after collection if shipping the same day is
not possible. Ensure that sufficient dry ice is used to keep the samples cold for
48 hours. Do not ship on a Friday or the samples will be ruined by the time we
get them on Monday.
For leaf tissue, use only 1ml 96-Deepwell plates from bioexpress.com or VWR:
From Bioexpress (www.bioexpress.com)
1 ml Deep-Well Riplate, Polypropylene, Non-Sterile, 20/case, #P-8700-1. (NOTE:
Use Beckman mats. Does NOT work with VWR mats.)
From Beckman (www.beckmancoulter.com)
Cap Mats for 1ml 96 well plate, Nonsterile, Case of 12, #267002 (NOTE: Use these
for the plates from Bioexpress, DO NOT USE BECKMAN PLATES, THEY
LEAK!)
From VWR (www.vwrsp.com)
1ml Polypropylene Deep Well Microplates, Nonsterile, Case of 50, #40002-009
Mat Lids For 1.0 mL Microplates, Nonsterile, Case of 50, #40002-001
KS USDA Sample Collection and Submission Protocol For Oven-Dried Fresh
Tissue
1. Always contact Guihua or Paul BEFORE collecting samples so that the
appropriate sample amounts and methods are used.
2. Always contact our lab BEFORE sending samples so that we can make sure
they arrive safely and are handled properly and promptly.
3. Always collect samples into approved 96-well plates.
4. We always work with 96-well sample plates. If you have fewer than 96
samples, you should replicate them to make better use of the sample plate.
5. Always leave the last well (H12) EMPTY so we can include 1 internal control
sample.
6. Always email a sample list in an excel file including: sample plate name and
number, well positions for each sample, sample name, traits of interest,
markers to use for each trait. Other info is helpful, such as sample generation,
pedigree, expected genotype for each marker (positive, negative, segregating,
etc.). See the Sample Excel file.
7. Always include samples of parent lines, known positives, or other controls.
8. Replication of parents or controls is helpful.
9. If you will be requesting newly published markers or non-published markers,
send the protocols 1 or 2 months prior to the samples.
10. Always send a UNIFORM amount of sample across wells.
11. Always label plates clearly with the plate name or number and the job name.
12. Collect 4 to 6 leaves (50 to 200mg total) of 2 cm length each (longer pieces do
not grind as well). Use only very young tissue, the 3 leaf stage works well. Do
not put more than 200mg of fresh tissue into a well.
13. Place plates WITHOUT LIDS into an oven at 38°C to 42°C, for 2 to 4 days. The
oven should have good air circulation and must NOT be above 48°C or DNA
degradation will occur. Leaves should remain green and snap when flexed.
14. Cap plates tightly and ship without ice. Plates MUST remain capped to
prevent re-hydration.
For leaf tissue, use only 1ml 96-Deepwell plates from bioexpress.com or VWR:
From Bioexpress (www.bioexpress.com)
1 ml Deep-Well Riplate, Polypropylene, Non-Sterile, 20/case, #P-8700-1. (NOTE:
Use Beckman mats. Does NOT work with VWR mats.)
From Beckman (www.beckmancoulter.com)
Cap Mats for 1ml 96 well plate, Nonsterile, Case of 12, #267002 (NOTE: Use these
for the plates from Bioexpress, DO NOT USE BECKMAN PLATES, THEY
LEAK!)
From VWR (www.vwrsp.com)
1ml Polypropylene Deep Well Microplates, Nonsterile, Case of 50, #40002-009
Mat Lids For 1.0 mL Microplates, Nonsterile, Case of 50, #40002-001