GB/T 37115-2018Protocol of diagnosis for infection with abalone herpersvirus (English PDF)

Foreword

This standard was drafted in accordance with the rules given in GB/T 1.1-2009.

1 Scope

This standard gives the clinical symptoms of alopecia virus disease, specifies the histopathological detection method of alopecia virus disease, real-time fluorescence polymerization Enzyme chain reaction detection method, polymerase chain reaction detection method.

This standard applies to the epidemiological investigation, monitoring, diagnosis and quarantine of alopecia virus disease.

2 Normative references

The following documents are indispensable for the application of this document. For dated references, only dated versions apply to this document. For undated references, the latest edition (including all amendments) applies to this document.

GB/T 6682 Analytical laboratory water specifications and test methods

GB/T 18088 entry and exit animal quarantine sampling

SC/T 7103 Technical specification for quarantine sampling of aquatic animals

3 Abbreviations

The following abbreviations apply to this document.

AbHV. Abalone Herpesvirus

CTAB. CetyltrimethylAmmonium Bromide

Ct value. Cycle threshold (Cyclethreshold)

EB. Ethidium Bromide

EDTA. Ethylene Diamine Tetraacetic Acid

PCR. Polymerase Chain Reaction (PolymeraseChainReaction)

qPCR. real-time fluorescent polymerase chain reaction (quantitativereal-timePolymeraseChainReaction)

4 Reagents and materials

4.1 Unless otherwise stated, the biochemical reagents used in this standard are analytically pure reagents.

4.2 If not specified, the test water should meet the specifications of GB/T 6682 water.

4.3 Taq enzyme and 10 times Taq enzyme concentration buffer; Taq enzyme concentration is 5U/µL, stored at -20 °C, to avoid repeated freezing and thawing.

4.4 dNTPs. containing 10 cmoL/L of dCTP, dGTP, dATP and dTTP.

4.5 qPCR primers and probes.

AbHVORF66F1. 5'-CCCGGACACCAGTAAGAAC-3'

AbHVORF66R1. 5'-AGGCTGCTATGCGTATGA-3'

Probe (AbHV66Prb1). 5'-6FAM-TGGCCGTCGAGATGTCCATG-TAMRA-3'

A 146 bp fragment of the ORF66 gene was amplified for fluorescent PCR. (See Appendix A)