GB/T 28062-2025Code of practice for detection of Candidatus Liberibacter asiaticus by quantitative real-time PCR (English PDF)

Foreword

This document is in accordance with the provisions of GB/T 1.1-2020 "Guidelines for standardization work Part 1: Structure and drafting rules for standardization documents" Drafting.

This document replaces GB/T 28062-2011 "Real-time fluorescence PCR detection method for citrus Huanglongbing bacteria" and is compatible with GB/T 28062-2011 Compared with the previous version, in addition to structural adjustments and editorial changes, the main technical changes are as follows.

— Changed the sampling method for Huanglongbing samples (see 9.1, 8.2 of the 2011 edition);

— Changed the Ct value of the positive control (see 11.3.2, 10.2.2 of the 2011 edition);

— Change the fluorescent dye detection primers CQULasF04/CQULasR04 of Bacillus asiaticus to more specific primers GGTAG-3'), and added the fluorescent labeled probe HLBp (sequence. 5'-AGACGGGTGAGTAACGCG-3') (See 10.1, F.1.1 of the 2011 edition);

— Added real-time fluorescence PCR detection of the target Bacillus asiaticus ribonucleotide reductase beta gene (nrdB gene) (See 11.3.3).

Please note that some of the contents of this document may involve patents. The issuing organization of this document does not assume the responsibility for identifying patents.

This document is proposed and coordinated by the National Technical Committee on Plant Quarantine Standardization (SAC/TC271).

This document was drafted by: National Agricultural Technology Extension Service Center, South China Agricultural University, and Southwest University Citrus Research Institute.

The main drafters of this document are Feng Xiaodong, Chen Ranran, Deng Xiaoling, Zheng Zheng, Wang Xuefeng and Zhou Changyong.

The previous versions of this document and the documents it replaces are as follows.

— First published in 2011 as GB/T 28062-2011;

— This is the first revision.

Detection of Citrus Huanglongbing Disease by Real-time Fluorescence Quantitative PCR Technical regulations

1 Scope

The quantitative PCR detection procedure specifies the laboratory setting, sample collection and processing, detection operation method and result judgment, and describes the confirmation method.

This document is applicable to the real-time fluorescence quantitative PCR of Bacillus asiaticus species in host plants and vectors of Huanglongbing (Psyllid citri citri) Detection and disease identification.

2 Normative references

GB 5040

GB/T 6682

3 Terms and definitions

The following terms and definitions apply to this document.

4 Principles of the method

The real-time fluorescence quantitative PCR detection used in this document includes two detection methods. fluorescent dye method and fluorescent probe method.

The method is to design primers using the unique DNA sequence of Huanglongbing fungus, add fluorescent dyes during PCR amplification, and the fluorescent dyes react with the products formed by amplification.