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GB/T 22333-2008: Reverse transcription polymerase chain reaction for Japanese B encephalitis virus
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| GB/T 22333-2008 | English | 239 |
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Reverse transcription polymerase chain reaction for Japanese B encephalitis virus
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GB/T 22333-2008
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Basic data | Standard ID | GB/T 22333-2008 (GB/T22333-2008) | | Description (Translated English) | Reverse transcription polymerase chain reaction for Japanese B encephalitis virus | | Sector / Industry | National Standard (Recommended) | | Classification of Chinese Standard | B41 | | Classification of International Standard | 11.220 | | Word Count Estimation | 6,640 | | Date of Issue | 2008-08-22 | | Date of Implementation | 2008-12-01 | | Regulation (derived from) | National Standard Approval Announcement 2008 No.14 (Total No.127) | | Issuing agency(ies) | General Administration of Quality Supervision, Inspection and Quarantine of the People's Republic of China, Standardization Administration of the People's Republic of China | | Summary | This standard specifies the technical requirements for reverse transcription -polymerase chain reaction test of Japanese encephalitis virus nucleic acid. This standard is applicable to the detection of animal brain tissue of Japanese B encephalitis virus nucleic acid. | GB/T 22333-2008: Reverse transcription polymerase chain reaction for Japanese B encephalitis virus
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Reverse transcription polymerase chain reaction for Japanese B encephalitis virus
ICS 11.220
B41
National Standards of People's Republic of China
Japanese encephalitis virus
Reverse transcription polymerase chain reaction test method
Posted 2008-08-22
2008-12-01 implementation
Administration of Quality Supervision, Inspection and Quarantine of People's Republic of China
Standardization Administration of China released
Foreword
Appendix A of this standard is a normative appendix.
The standard proposed by the People's Republic of China Ministry of Agriculture.
This standard by the National Standardization Technical Committee on Animal Epidemic Prevention.
This standard was drafted. the Ministry of Agriculture veterinary diagnostic center.
The main drafters of this standard. Wang Hongwei, Sun Ming, Wang Chuanbin, Zhao Chen Xi, TIAN Ke-gong.
Japanese encephalitis virus
Reverse transcription polymerase chain reaction test method
1 Scope
This standard specifies the technical requirements of reverse transcription polymerase chain reaction test of Japanese encephalitis virus nucleic acid.
This standard is applicable to the detection of animal brain tissue Japanese encephalitis virus nucleic acid.
2 Reagents
2.1 denaturing solution (see Section A.1 chapter).
2.2 2mol/L sodium acetate (pH4.0).
2.3 phenol - chloroform - isoamyl alcohol mixture (see Section A.2 chapter).
2.4 2.5mmol/LdNTP.
2.5 8mmol/L on the downstream primer mixture (primer sequences see section A.3 chapter).
2.6 10x PCR buffer.
2.7 1% ethidium bromide (EB) solution.
2.8 TAE electrophoresis buffer.
2.9 1% agarose gel.
2.10 loading buffer (see Chapter A.4).
2.11 Other reagents. 10U/μLAMV reverse transcriptase, 100bpDNA molecular mass standard, 50U/μLRNA inhibitors,
5U/μLT Rao q DNA polymerase, isopropanol, 75% ethanol solution, 25mmol L magnesium chloride solution /, DEPC-treated sterile double
Distilled water.
3 laboratory conditions
3.1 laboratory level requirements
Laboratories must biosafety level Ⅲ (BSL-3 level) laboratories.
3.2 Laboratory partition
RT-PCR test-area reaction liquid preparation area (dosing area), nucleic acid extraction zone, the amplified region, zone electrophoresis. Flow sequence for the dosing area →
Amplified nucleic acid extraction District → District → electrophoresis area. Prohibited equipment and reagents back.
3.3 laboratory should be equipped with equipment and supplies
3.3.1 Instrument. analytical balance, high-speed centrifuge, a vacuum dryer, PCR amplification device, electrophoresis, electrophoresis tank, UV gel imager (or purple
Outside the analyzer), liquid nitrogen tanks or -70 ℃ refrigerator, microwave, tissue grinder, -20 ℃ refrigerator, water bath, adjustable pipette (maximum range points
Do as 2μL, 20μL, 200μL, 1000μL).
3.3.2 supplies. scissors ophthalmology, ophthalmic tweezers, weighing paper, 10mL disposable syringe, 1.5mL sterile centrifuge tube, 0.2mL thin wall PCR
Pipe, agarose, 500mL graduated cylinder, 500mL conical flask, pipette tips (10μL, 200μL, 1000μL), sterilized double-distilled water.
Procedure 4
Sample collection and processing 4.1
4.1.1 Sample collection. the dying animals, culling of animals or aborted fetus brain tissues.
4.1.2 Sample treatment. The samples were each treated.
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