Severe acute respiratory syndrome (SARS) was first reported in Asia in February 2003, and then spread to more than a dozen countries around the world. SARS is a type of coronavirus that has a capsule and a linear single-stranded genome. The corresponding structure diagram of the RNA virus is as follows:

According to the introduction of CDC and WHO, nucleic acid diagnosis for SARS is generally for ORF1ab/RdRP (RNA-dependent RNA polymerase), Gene N, Gene M, and Gene E. Gene S is more used for the development of corresponding drugs and vaccines. The sequence included in NCBI is NC_004718.3, with a total length of 29751bp, and the gene encodes 4 structural proteins: S, M, N and E.

The detection limit of the kit is the biggest cause of false negatives. Therefore, for the detection limit of diagnostic products, we need to carefully evaluate the performance of each kit. In the past, performance evaluation for detection limits generally selected in vitro transcribed RNA or clinically positive viruses as standard products for performance evaluation. However, both have obvious certainty. First, the RNA transcribed in vitro is generally very pure and single, does not involve extraction, and does not meet the microenvironment of the clinical virus. Therefore, the detection sensitivity will inevitably be overestimated. This is also a high probability of false The main reason for the negative; clinically positive virus again, even if it is an inactivated virus, still has a great biological safety hazard. Therefore, the laboratory level is required to be P3-P4. Most of the units that develop diagnostic kits do not meet the requirements. The source of clinically positive viruses is also very limited and cannot be stably supplied, and is repeatedly used for performance evaluation of the kit.

It has the complete envelope structure of the virus and the nucleic acid sequence corresponding to SARS, but the pseudovirus standard product with low biosafety risk perfectly overcomes the shortcomings of choosing in vitro transcribed RNA or clinically positive virus as the standard product. It is very suitable Standards for evaluation of nucleic acid diagnostic performance.

Construction process: SARS gene synthesis → pseudovirus packaging → purification → QC detection (ddPCR)

ORF1ab/RdRP（2795bp）、Gene N（1207bp）

·The SARS pseudovirus standard product is different from the RNA synthesized in vitro and clinically positive live virus. It perfectly overcomes the shortcomings of both, and is really suitable for performance evaluation of the kit, including detection limit, specificity, repeatability, etc.;

·According to the characteristics of SARS coronavirus, a total of 2 segments of ORF1ab/RdRP (RNA-dependent RNA polymerase) and Gene N (nucleocapsid phosphoprotein) were selected for the packaging of pseudoviruses, which well characterized the characteristics of SARS sequence;

·Coby Bio has designed the primers and probe sequences of the ddPCR system, which can complete the QC copy number detection of pseudovirus without constructing a standard curve, which can very accurately characterize the copy number of the standard product, and overcome the Q-PCR analysis of CT Inaccuracy of the relative judgment standard of the value;

·Pseudovirus standard products, non-pathogenic, renewable, reliable quality control methods, stable between batches, can be prepared and supplied stably for a long time, and require laboratory biosafety level P2 to meet the safety needs of many organizations.

ORF1ab/RdRP:

TCTGCGGATGCATCAACGTTTTTAAACGGGTTTGCGGTGTAAGTGCAGCCCGTCTTACACCGTGCGGCACAGGCACTAGTACTGATGTCGTCTACAGGGCTTTTGATATTTACAACGAAAAAGTTGCT

GGTTTTGCAAAGTTCCTAAAAACTAATTGCTGTCGCTTCCAGGAGAAGGATGAGGAAGGCAATTTATTAGACTCTTACTTTGTAGTTAAGAGGCATACTATGTCTAACTACCAACATGAAGAGACTAT

TTATAACTTGGTTAAAGATTGTCCAGCGGTTGCTGTCCATGACTTTTTCAAGTTTAGAGTAGATGGTGACATGGTACCACATATATCACGTCAGCGTCTAACTAAATACACAATGGCTGATTTAGTCT

ATGCTCTACGTCATTTTGATGAGGGTAATTGTGATACATTAAAAGAAATACTCGTCACATACAATTGCTGTGATGATGATTATTTCAATAAGAAGGATTGGTATGACTTCGTAGAGAATCCTGACATC

TTACGCGTATATGCTAACTTAGGTGAGCGTGTACGCCAATCATTATTAAAGACTGTACAATTCTGCGATGCTATGCGTGATGCAGGCATTGTAGGCGTACTGACATTAGATAATCAGGATCTTAATGG

GAACTGGTACGATTTCGGTGATTTCGTACAAGTAGCACCAGGCTGCGGAGTTCCTATTGTGGATTCATATTACTCATTGCTGATGCCCATCCTCACTTTGACTAGGGCATTGGCTGCTGAGTCCCATA

TGGATGCTGATCTCGCAAAACCACTTATTAAGTGGGATTTGCTGAAATATGATTTTACGGAAGAGAGACTTTGTCTCTTCGACCGTTATTTTAAATATTGGGACCAGACATACCATCCCAATTGTATT

AACTGTTTGGATGATAGGTGTATCCTTCATTGTGCAAACTTTAATGTGTTATTTTCTACTGTGTTTCCACCTACAAGTTTTGGACCACTAGTAAGAAAAATATTTGTAGATGGTGTTCCTTTTGTTGT

TTCAACTGGATACCATTTTCGTGAGTTAGGAGTCGTACATAATCAGGATGTAAACTTACATAGCTCGCGTCTCAGTTTCAAGGAACTTTTAGTGTATGCTGCTGATCCAGCTATGCATGCAGCTTCTG

GCAATTTATTGCTAGATAAACGCACTACATGCTTTTCAGTAGCTGCACTAACAAACAATGTTGCTTTTCAAACTGTCAAACCCGGTAATTTTAATAAAGACTTTTATGACTTTGCTGTGTCTAAAGGT

TTCTTTAAGGAAGGAAGTTCTGTTGAACTAAAACACTTCTTCTTTGCTCAGGATGGCAACGCTGCTATCAGTGATTATGACTATTATCGTTATAATCTGCCAACAATGTGTGATATCAGACAACTCCT

ATTCGTAGTTGAAGTTGTTGATAAATACTTTGATTGTTACGATGGTGGCTGTATTAATGCCAACCAAGTAATCGTTAACAATCTGGATAAATCAGCTGGTTTCCCATTTAATAAATGGGGTAAGGCTA

GACTTTATTATGACTCAATGAGTTATGAGGATCAAGATGCACTTTTCGCGTATACTAAGCGTAATGTCATCCCTACTATAACTCAAATGAATCTTAAGTATGCCATTAGTGCAAAGAATAGAGCTCGC

ACCGTAGCTGGTGTCTCTATCTGTAGTACTATGACAAATAGACAGTTTCATCAGAAATTATTGAAGTCAATAGCCGCCACTAGAGGAGCTACTGTGGTAATTGGAACAAGCAAGTTTTACGGTGGCTG

GCATAATATGTTAAAAACTGTTTACAGTGATGTAGAAACTCCACACCTTATGGGTTGGGATTATCCAAAATGTGACAGAGCCATGCCTAACATGCTTAGGATAATGGCCTCTCTTGTTCTTGCTCGCA

AACATAACACTTGCTGTAACTTATCACACCGTTTCTACAGGTTAGCTAACGAGTGTGCGCAAGTATTAAGTGAGATGGTCATGTGTGGCGGCTCACTATATGTTAAACCAGGTGGAACATCATCCGGT

GATGCTACAACTGCTTATGCTAATAGTGTCTTTAACATTTGTCAAGCTGTTACAGCCAATGTAAATGCACTTCTTTCAACTGATGGTAATAAGATAGCTGACAAGTATGTCCGCAATCTACAACACAG

GCTCTATGAGTGTCTCTATAGAAATAGGGATGTTGATCATGAATTCGTGGATGAGTTTTACGCTTACCTGCGTAAACATTTCTCCATGATGATTCTTTCTGATGATGCCGTTGTGTGCTATAACAGTA

ACTATGCGGCTCAAGGTTTAGTAGCTAGCATTAAGAACTTTAAGGCAGTTCTTTATTATCAAAATAATGTGTTCATGTCTGAGGCAAAATGTTGGACTGAGACTGACCTTACTAAAGGACCTCACGAA

TTTTGCTCACAGCATACAATGCTAGTTAAACAAGGAGATGATTACGTGTACCTGCCTTACCCAGATCCATCAAGAATATTAGGCGCAGGCTGTTTTGTCGATGATATTGTCAAAACAGATGGTACACT

TATGATTGAAAGGTTCGTGTCACTGGCTATTGATGCTTACCCACTTACAAAACATCCTAATCAGGAGTATGCTGATGTCTTTCACTTGTATTTACAATACATTAGAAAGTTACATGATGAGCTTACTG

GCCACATGTTGGACATGTATTCCGTAATGCTAACTAATGATAACACCTCACGGTACTGGGAACCTGAGTTTTATGAGGCTATGTACACACCACATACAGTCTTGCAG

Gene N:

ATGTCTGATAATGGACCCCAATCAAACCAACGTAGTGCCCCCCGCATTACATTTGGTGGACCCACAGATTCAACTGACAATAACCAGAATGGAGGACGCAATGGGGCAAGGCCAAAACAGCGCCGACC

CCAAGGTTTACCCAATAATACTGCGTCTTGGTTCACAGCTCTCACTCAGCATGGCAAGGAGGAACTTAGATTCCCTCGAGGCCAGGGCGTTCCAATCAACACCAATAGTGGTCCAGATGACCAAATTG

GCTACTACCGAAGAGCTACCCGACGAGTTCGTGGTGGTGACGGCAAAATGAAAGAGCTCAGCCCCAGATGGTACTTCTATTACCTAGGAACTGGCCCAGAAGCTTCACTTCCCTACGGCGCTAACAAA

GAAGGCATCGTATGGGTTGCAACTGAGGGAGCCTTGAATACACCCAAAGACCACATTGGCACCCGCAATCCTAATAACAATGCTGCCACCGTGCTACAACTTCCTCAAGGAACAACATTGCCAAAAGG

CTTCTACGCAGAGGGAAGCAGAGGCGGCAGTCAAGCCTCTTCTCGCTCCTCATCACGTAGTCGCGGTAATTCAAGAAATTCAACTCCTGGCAGCAGTAGGGGAAATTCTCCTGCTCGAATGGCTAGCG

GAGGTGGTGAAACTGCCCTCGCGCTATTGCTGCTAGACAGATTGAACCAGCTTGAGAGCAAAGTTTCTGGTAAAGGCCAACAACAACAAGGCCAAACTGTCACTAAGAAATCTGCTGCTGAGGCATCT

AAAAAGCCTCGCCAAAAACGTACTGCCACAAAACAGTACAACGTCACTCAAGCATTTGGGAGACGTGGTCCAGAACAAACCCAAGGAAATTTCGGGGACCAAGACCTAATCAGACAAGGAACTGATTA

CAAACATTGGCCGCAAATTGCACAATTTGCTCCAAGTGCCTCTGCATTCTTTGGAATGTCACGCATTGGCATGGAAGTCACACCTTCGGGAACATGGCTGACTTATCATGGAGCCATTAAATTGGATG

ACAAAGATCCACAATTCAAAGACAACGTCATACTGCTGAACAAGCACATTGACGCATACAAAACATTCCCACCAACAGAGCCTAAAAAGGACAAAAAGAAAAAGACTGATGAAGCTCAGCCTTTGCCG

CAGAGACAAAAGAAGCAGCCCACTGTGACTCTTCTTCCTGCGGCTGACATGGATGATTTCTCCAGACAACTTCAAAATTCCATGAGTGGAGCTTCTGCTGATTCAACTCAGGCATAA