GB/T 25165-2010Protocol of identification of bovine, caprine, ovine and porcine derived materials in gelatin - Real time PCR method (English PDF)
明胶中牛、羊、猪源性成分的定性检测方法 实时荧光PCR法
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Issued by
SAMR; SAC
Level / Type
National · Recommended
Issue date
September 26, 2010
Implementation date
May 1, 2011
Scope
GB/T 25165-2010 is the English-translated version of 明胶中牛、羊、猪源性成分的定性检测方法 实时荧光PCR法.
GB/T 25165-2010 is the Chinese standard "Protocol of identification of bovine, caprine, ovine and porcine derived materials in gelatin -- Real time PCR method".
Its scope clause reads: This Standard specifies the real time PCR detection method of bovine, ovine and porcine derived components in gelatin. This Standard applies to the qualitative identification of bovine, ovine and porcine derived components in gelatin.
The detection limit of this method is 0.1%. Its clauses include terms and definitions; principle; primers and probes for detection; reagents; solution preparation; instruments; sample collection; inspection steps; result determination; measures to prevent cross-contamination during detection.
It was issued by the General Administration of Quality Supervision, Inspection and Quarantine on SEPTEMBER 26, 2010 and took effect on MAY 01, 2011.
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Document preview — GB/T 25165-2010
National Standard of the People's Republic of China
- ICS
- 65.020.30
- Classification
- B 43
Issued by: State Administration for Market Regulation; Standardization Administration of the PRC
Contents
- Foreword3
- 1 Scope4
- 2 Normative references4
- 3 Terms and definitions4
- 4 Principle5
- 5 Primers and probes for detection5
- 6 Reagents5
- 7 Solution preparation6
- 8 Instruments6
- 9 Sample collection6
- 10 Inspection steps6
- 11 Result determination8
- 12 Measures to prevent cross-contamination during detection8
1 Scope
This Standard specifies the real time PCR detection method of bovine, ovine and porcine derived components in gelatin.
This Standard applies to the qualitative identification of bovine, ovine and porcine derived components in gelatin. The detection limit of this method is 0.1%.
2 Normative references
The provisions in following documents become the provisions of this Standard through reference in this Standard. For dated references, the subsequent amendments (excluding corrigendum) or revisions do not apply to this Standard, however, parties who reach an agreement based on this Standard are encouraged to study if the latest versions of these documents are applicable. For undated references, the latest edition of the referenced document applies.
GB/T 6682, Water for analytical laboratory use - Specification and test methods
GB 6783, National Food Safety Standard Food additive - Gelatine
GB/T 14699.1, Feeding Stuffs - Sampling
GB/T 27403-2008, Criterion on quality control of laboratories - Molecular biological testing of food
3 Terms and definitions
For the purposes of this document, the following terms and definitions apply.
3.1 Real time PCR
add fluorescent groups to the PCR reaction system, and use the accumulation of fluorescent signals to monitor the entire PCR amplification process in real time
3.2 Cycle threshold
meets the requirements of secondary water in GB/T 6682.
7 Solution preparation
7.1 Lysate
Ingredients include: 2% (mass concentration) CTAB (cetyltrithylammonium bromide, cetyltrimethylammonium bromide), 100mmol/L Tris (tris hydroxymethyl aminomethane, trishydroxymethyl aminomethane), 1.4 mol/L NaCl, 20mmol/L EDTA (ethylenediaminetetraacetic acid, ethylenediaminetetraacetic acid). Use HCl to adjust pH to 8.0.
7.2 Real time PCR reaction mix
The 12.5µL reaction system includes: 1U~2U (Unit, enzymatic unit) of Taq enzyme, 1×PCRbuffer, 2.5mmol/L~4.0mmol/L Mg2+, 0.2U~1U UNG enzyme, 0.2mmol/L d (A,C,G) TPs, 0.2mmol/L~0.4mmol/L dUTP, 400nmol/L ROX dye (some fluorescent PCR instruments do not require ROX correction).
8 Instruments
8.1 Real time PCR instrument.
8.2 Constant temperature water bath.
8.3 Centrifuge: the centrifugal force is not less than 3000g.
8.4 Micropipette: 0.5µL~10µL, 10µL~100µL, 20µL~200µL, 200µL~1000µL.
8.5 Nucleic acid protein analyzer or UV spectrophotometer.
8.6 pH meter.
8.7 Balance: the resolution is 0.01g.
9 Sample collection
Conduct sampling of edible gelatin according to GB 6783. Sample feed gelatin according to GB/T 14699.1. Pulverize the experimental sample. Mix well and set aside. […]
10 Inspection steps
10.1 DNA extraction
......
This preview omits tables, figures, formulas and parts of the technical clauses. The complete document — 15 pages — is available in the English PDF.
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Related Standards
GB 6783-2013 — Food additive -- Gelatin
GB/T 14699.1-2005 — Feeding stuffs -- Sampling
GB/T 27403-2008 — Criterion on quality control of laboratories -- Molecular biological testing of food
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