T/GDSF 0003-2023 罗非鱼无乳链球菌和海豚链球菌双重荧光PCR检测方法

ICS65.150

CCSB50

GDSF

广东水产学会团体标准

T/GDSF0003—2023

罗非鱼无乳链球菌和海豚链球菌双重荧光

PCR检测方法

DuplexReal-timePCRMethodfortheDetectionofStreptococcusagalactiaeand

StreptococcusiniaeinTilapia

2023-9-4发布2023-9-5实施

广东水产学会发布

T/GDSF0003—2023

目次

前言................................................................................III

1范围................................................................................1

2规范性引用文件......................................................................1

3术语和定义..........................................................................1

4缩略语..............................................................................1

5试剂和耗材..........................................................................1

试剂..............................................................................1

引物和探针........................................................................1

6仪器设备............................................................................2

7样品采集与处理......................................................................2

样品采集..........................................................................2

7.1.1采样工具........................................................................2

7.1.2待检样本........................................................................2

7.1.3样品运输........................................................................2

样品前处理........................................................................2

7.2.1池塘水样品前处理................................................................2

7.2.2环境拭子前处理..................................................................2

7.2.3细菌培养物......................................................................2

7.2.4组织样品前处理..................................................................2

8操作方法............................................................................2

DNA提取...........................................................................3

反应体系的配制....................................................................3

加样..............................................................................3

荧光PCR反应......................................................................3

8.4.1上机............................................................................3

8.4.2循环条件设置....................................................................3

8.4.3荧光通道设定....................................................................3

9结果判定............................................................................3

阈值设定..........................................................................3

质控标准..........................................................................3

结果描述及判定....................................................................3

附录A（规范性）溶液配置........................................................5

A.10.01mol/LPBS(pH7.2)溶液的配置..................................................5

A.2阳性对照..........................................................................5

A.3阴性对照..........................................................................5

附录B（规范性）引物、探针的名称、序列和工作浓度................................6

I