Custom cDNA Synthesis

BOC Sciences offers cDNA synthesis that uses RNA as a template to obtain cDNA by reverse transcription. The cDNA can then be used as a template for various downstream applications in RNA research.

What is cDNA?

cDNA (complementary DNA) is synthetic DNA whose base sequence is complementary to that of DNA. cDNA is synthesized by the action of RNA-dependent DNA polymerase (reverse transcriptase) in the presence of appropriate primers using RNA as a template, and after synthesizing a single-stranded cDNA and then treating it with a base to remove its corresponding RNA, it is then synthesized by the action of DNA-dependent DNA polymerase using the single-stranded cDNA as a template. After single-stranded cDNA is synthesized and its corresponding RNA is removed by alkali treatment, single-stranded cDNA is used as a template to synthesize double-stranded cDNA by the action of DNA-dependent or RNA-dependent DNA polymerase.

Procedure for cDNA synthesisFig.1 Procedure for cDNA synthesis. (Meis et al., 2009)

BOC Sciences' cDNA Synthesis Service

BOC Sciences provides cDNA synthesis services in which the synthesized first-strand cDNA can be widely used for second-strand cDNA synthesis for PCR amplification, hybridization, cDNA library creation, and full-length cDNA synthesis.

  • First-strand cDNA Synthesis
    First-strand cDNA synthesis utilizes oligo (dT), random primers, or a combination of these strategies to initiate the reverse transcription reaction. Initiation of the reaction with oligo(dT) preferentially starts at the 3' end of the RNA fragment.
  • Second-strand cDNA Synthesis
    Methods for synthesizing the second-strand of cDNA can be selected depending on the needs and application. The methods we currently offer include the following.
Self-priming methodThe 3' end of the synthesized single-stranded cDNA is able to form a short hairpin structure, which provides a ready-made primer for the synthesis of the second strand. The second strand of the cDNA is synthesized by using the E. coli DNA polymerase I Klenow fragment or reverse transcriptase after denaturation of the DNA-RNA hybridization strand from the product of the first-stranded synthesis reaction, and the loop is digested with S1 nuclease specific for the single strand, which can be further cloned.
Replacement synthesis methodIn the presence of dNTP, RNAase H is utilized to create cuts and nicks in the mRNA strand of the hybridized strand. This results in a series of RNA primers for the synthesis of the second strand, which is synthesized under the action of E. coli DNA polymerase I.
  • Analysis of cDNA
    The cDNA synthesis product needs to be validated and analyzed to ensure that the customer gets the target product.
Analysis methodSpecifications
Gel ElectrophoresisThe size and purity of the cDNA can be initially determined by the electrophoretic migration rate.
Real-time quantitative PCR (qPCR)qPCR measures the expression level of a target gene and provides relative or absolute quantitative results.
Sequence AnalysisSanger sequencing or high-throughput sequencing technology enables cDNA sequence analysis.

cDNA Synthesis Steps

  • Sample Preparation
    RNA is the template for cDNA synthesis, and RNA needs to maintain its integrity during the extraction process.
  • Removal of Genomic DNA
    Before cDNA synthesis, DNase I must be completely removed because the residual enzyme degrades single-stranded DNA. Double-stranded specific DNase can be used as an alternative to DNase I to remove contaminating gDNA without affecting RNA or single-stranded DNA.
  • Selecting a Reverse Transcriptase
    Depending on the need and application, select the appropriate reverse transcriptase to ensure efficient cDNA synthesis.
  • Formulating the Reaction System
    In addition to the enzyme and primers, the main reaction components for reverse transcription include an RNA template (pre-treated to remove genomic DNA), buffer, dNTP, DTT, RNAase inhibitor and nuclease-free water.
  • cDNA Synthesis
    cDNA synthesis includes cDNA first and second-strand synthesis.

Materials to be Provided by the Customer

  • Total RNA: volume ≥10μL, concentration ≥50ng/μL
  • Cell sample: 106 cells or more, cell suspension or cell precipitation
  • Tissue sample: volume of tissue ≥100mg


  1. Meis J E, et al. RNA amplification and cDNA synthesis for qRT-PCR directly from a single cell[J]. 2009.
* Only for research. Not suitable for any diagnostic or therapeutic use.
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