Questions and Answers 24 July.pdf
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- Attached to
- mRNA-sequencing and Methyl-Bisulfite sequencing from submitted leaf tissue samples Federal contract opportunity
- Solicitation number
- 1232SA26Q0964
About this file
This is a Questions and Answers (Q&A) document clarifying requirements for a genomic sequencing contract involving cereal crop plant samples.
The contract involves RNA sequencing and whole genome bisulfite sequencing (WGBS) services for leaf samples from three cereal crop species: Wheat (14 Gb genome), Barley (5 Gb genome), and Oats (10-12 Gb genome). The client will submit a maximum of 610 samples for RNA analysis and 20 samples for methylation analysis over a one-year performance period, with samples arriving on a rolling basis with no minimum batch size. RNA sequencing shipments typically contain 20-100 samples, while methylation samples range from 4-20 samples per shipment. All work should be performed at a single facility if cost-effective. For WGBS analysis, the client requires 20X coverage targeting Barley or Oats samples. The contractor will receive samples in various formats—plant tissue, DNA, or RNA—based on what individual researchers submit, and should accommodate vendor requirements for tissue preparation and quantity. Quality control reports are expected within 3-6 business days of sample receipt, regardless of whether samples arrive as plant tissue or extracted DNA/RNA. The client will perform its own genome assembly and alignment using reference genomes (Morex V3 for Barley, OT3098v2 for Oats, Csv2.1 for Wheat) and requires only raw sequencing data without assembly or alignment services. The client is open to alternative methylation library options beyond bisulfite treatment.
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Text version
1. Can you confirm the exact species (or species per CLIN, if they differ) for the leaf samples?
These are Cereal Crops plant samples. They can be Wheat, Barley or Oats
2. What is the reference genome size or size range for the species involved? This is especially critical for accurately quoting the WGBS project (CLIN 0002).
Wheat -14Gb, Barley- 5Gb and Oats -10-12 Gb
3. Will samples be sent to 1 facility or is your client open to sending them to different facilities for RNA sequencing and WGBS if it's more cost-effective?
Ideally, we want all of them to go to one facility.
4. For CLIN 0002 (WGBS), what sequencing depth/coverage are you targeting per sample?
We would like to see a 20X coverage and these samples are either in Barley or Oats
5. Can you clarify what "assembled data" means for each CLIN (RNA-Seq vs. WGBS) — e.g., reference alignment, expression/methylation calling, or de novo assembly?
We think assembled data means reference alignment. For this purpose, raw data generated can be fine. We do not need assembling.
6. Is there a specific reference genome assembly/version you'd like used for alignment (if applicable)?
We have expertise to do the genome assembly. Hence we do not need them to do any assembly or alignment. For their reference, we use Morex V3 for Barley, OT3098v2 for Oats and Csv2.1 for Wheat.
7. How will the leaf tissue samples be submitted (e.g. flash frozen, in RNAlater, silica-dried, etc.)?
We can submit samples based on the Vendors requirements.
8. How much tissue will be submitted per sample?
We can submit tissue samples and quantities based on the Vendors requirements.
9. Since samples will ship on a rolling basis over the 1-year period of performance, can you please estimate the typical shipment size and frequency?
Typically, the average sample size will be above 20 and less than 100 samples for a RNAseq study and for methylation they can be 4-20 samples.
10. The SOW described the sample tissue will be submitted on an “as-needed” basis. Is there an anticipated minimum sample number that would be submitted at any single time, or should we assume that as little as 1 sample may be submitted in a batch?
There is no anticipated minimum sample size or number that will be submitted at any single time. The samples will be submitted by multiple people in the group under this contract.
11. The SOW notes a total quantity of 610 samples for RNA analysis and 20 samples for methyl analysis. Is this number the minimum or maximum potential number of samples through the period of performance?
This is maximum numbers of samples throughout the performance period.
12. The SOW technical description notes that total RNA or DNA would be submitted.
However, the Part Name/Number in the table and the deliverables details of the SOW also mentions. Can we please have clarification on the type of material that would be submitted?
The samples will be submitted as plant tissue, DNA, or RNA. For example, few people can submit samples as plant tissue and others who would like to submit DNA/RNA can submit accordingly.
13. Under “Key Deliverables” of the SOW, can you please confirm if the QC report of the submitted samples are 3 or 6 business days after submitted sample is received?
And would this assume submission of the RNA or DNA (excluding extraction)? And would this assume submission of the RNA or DNA (excluding extraction)?
The samples QC can be 3 - 6 business days. This generally depends on how quickly they can get to the samples. We are ok with a language that would be anywhere between those numbers. Since QC is done by the company getting into contract, they would be able to tell us whether samples are good or bad, whether it is plant tissue or DNA/RNA
14. Will the investigator be open to an alternative library option using enzymatic methylation, or is bisulfite treatment a requirement?
Yes, we can be open to other methylation library options.
15. The SOW does not specify the amount of data needed for the 20 Bisulfite sequencing. Can you specify?
We have an 11 Gb genome for oats. We are hoping to get 20x coverage for the genome for the methylation.
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