When developing an MDx (molecular diagnostic) assay, it is vital to use the proper reference standards and controls. Reference standards, which often mimic natural biological materials, are used for developing the assay itself, whereas controls, which are often fragments of biological materials, are used to assess final working assay quality. While there are many such predesigned and tested products available to purchase, customization allows flexibility not otherwise possible. Our Custom Molecular (DNA and RNA) Reference Standards & Controls are platform independent and ideal for use with common MDx assay technologies, such as PCR and NGS.
For example, if you are developing an RT-qPCR assay for the detection and quantification of an RNA virus involved in respiratory disease, such as SARS-CoV-2, any or all of the following might be needed:
*Whole viral genomes are not allowed for safety reasons
For anything marked as ‘Inquire’ or If you have needs that are different from the other general specifications presented, please fill out the Optional Information box on the request form.
After we receive your desired sequence, manufacturing of reference standards and controls occurs via the following steps (see Figure 1 for a condensed, graphical version of dsDNA and ssRNA manufacturing):
1. Synthesis of the dsDNA gene or fragment
2. Cloning of the gene or fragment into a plasmid
3. Sequencing verification of the plasmid insert
4. Synthesis of plasmid-specific PCR primers
5. PCR amplification of the cloned insert
6. Purification of PCR-amplified insert via spin column
7. QC of PCR-amplified insert via capillary electrophoresis
a) End of process for dsDNA product
8. In vitro transcription of PCR-amplified insert
9. Removal of DNA template with DNase
10. Purification of RNA via spin column
11. Quality control of RNA via capillary electrophoresis
a) End of process for ssRNA product
12. Separate syntheses of sense and antisense DNA strands
13. Pooling of both strands in equimolar amounts
14. In vitro transcription and hybridization
a) End of process for dsRNA product
Figure 1.Following cloning of the gene or fragment, PCR is used to amplify the desired insert. Direct PCR leads to dsDNA products. IVT (in vitro transcription) of the PCR product leads to the ssRNA product.