DB5132-T 88-2023 牦牛肉PCR一膜芯片技术检验方法

ICS

DB5132四川省（阿坝藏族羌族自治州）地方标准DB

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牦牛肉

PCR—膜芯片技术检验方法Identification

Yak

Meat

with

PCR-membrane

Chip

Technology 阿坝藏族羌族自治州市场监督管理局发布

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  本文件按照GB/T

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  III

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PCR-膜芯片技术检验方法

PCR

将预先制备的DNA探针有序的固定于尼龙膜表面，然后与标记的样品杂交，通过对杂交信号的检测

Sodium

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SSPE：核酸杂交缓冲液ddH2O：双重蒸馏水

针对牦牛肉及市场常见假冒牦牛肉动物源性成分特异基因片段设计引物，对DNA模板进行多重聚合

膜芯片杂交仪（MFS-8)

6mm

15×

去活化清洗液（2×SSPE，0.1%

杂交清洗液（2×SSPE，

mmol/L

mmol/EDTA·Na2，其

溴-4-氯

-3-

mmol/L

mmol/L

=

实验用水符合GB/

/

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PCR

表1

表1

PCR

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按GB/T

PCR

中提取并纯化后的DNA为模板，在

20μL

体系中加入

μL、正向引物（20

μ

μL、反向引物（20

μL、模板

μL，按以下条件进行扩增

延伸

个循环

cm的膜条，将裁好的尼龙膜放置于双蒸水中浸泡

μ

1）。图1

膜芯片去活化

：将制备好的膜芯片放入杂交盒内，芯片标签正面朝上，加入

去活

45℃，90

/min

-----+--PCR扩增失败，重复试验++--++-+注：“+”为阳性，“-”为阴性。

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洗膜

：吸除杂交体系液，加入

预热好的杂交清洗液