Establishment of National Genetically Modified Product Detection Laboratories and Detection

Method Complex

ZHU Shui-fang, Ph.D

GMOs detection center

Institute of Animal and Plant Quarantine

1. Detection technology of GMOs

Real-time PCR

DNA Microarray

Captured PCR-ELISA

Quicktest strip

2. Publications for GMOs detection

12 SN standard protocols

8 GB standard protocols

4 for regulation and detection technology

3. Construction of laboratories for GMOs detection

Research contents

Detection technology of GMOs

• Real-time PCR• DNA Microarray • Captured PCR-ELISA • Quicktest strip

Real-time PCRScreening Kits Target genes: CaMV 35S Promotor, Nos teminator , NptII, Bar, FMV promotor, Pat

Reference gene: 18S rRNA

Quantitative Kits

Roundup Ready soybean

Bt176 Maize

Event-specific detection kits GTS40-3-2, Bt176, Mon810, Bt11, GA21, T25, RT73

Real time PCR - TaqMan™ Technology

Positive sample

Negative sample

Ct value

Screening Detection of GM potato and rapeseed

GM Soy Analysis Std Curve

y = -1.5389Ln(x) +7.1327

R2 = 0.9981

0.00

2.00

4.00

6.00

8.00

10.00

12.00

0.01 0.10 1.00 10.00

% GM

?C

t Calculated StdCurve

Amplification of EPSPS gene Amplification of lectin gene

Quantitative detection of Roundup Ready Soybean

Real-time PCR kits

DNA Microarray

Total: 46 genes

31 inserted genes

e.g. EPSPS

7 reference genes

e.g. lectin

Genes involved in microarray research

8 event-specific genes

e.g. Roundup Ready Soybean

DNA Microarray Technology

Versatile chip

9 genes

DNA Microarray

Screening chip Event-specific chip

Species chip Comprehensive chip

GM Soybean, maize

rapeseed, cottonDetection of food and feed

Event-specific sequence

P35s, Pnos, NptII

CaMV35S!

Nos, FMV35S

Bar, Pat1(maize)

Pat2 (rapeseed)

Bt176, Bt11, Mon810, T14/25, GA21, CBH-351 maize

Bioscien toamto

Mon531 cotton

Multiple PCR and hybridization results of rapeseed

Multiple PCR and hybridization results of GM maize

Specific detection for RUR soybean

Specific detection for Mon810 maize

Multiple PCR and hybridization results of soybean

Multiple PCR and hybridization results of cotton

Specific detection for Bt11 maize

Specific detection for GA21 maize

Specific detection for Bt176 maize

DNA Microarray kits

Captured PCR-ELISA

Capture of target nucleic acid molecule

PCR-ELISA

Affinity capture of target DNA molecule

Adding sample Target DNA molecule

impurity molecule

washing

electrophoresis hybridization

probe

PCR product

PCR-ELISA Procedure

Quicktest strip for Cry1Ab/Ac

control

test

positive negative

Publications for GMOs detection

4 for regulation and detection technology

12 SN standard protocols

8 GB standard protocols

GMOs regulation and detection technology

2 for regulation, 2 for detection technology

SN and GB protocols for GMOs detection

12 SN (issued in 2003),

8 GB (will soon be issued)

Detection methods (revising, ISO 7th Working group )

Bioscien tomato, Mon531 cotton, RT73 rapeseed, Edible Oil

Publications of GMOs regulation and detection technology

12 SN standard protocols

•The general requirements for the laboratories for gene detection and identification

• Protocol of the Qualitative Polymerase Chain Reaction (PCR) for Detecting Genetically Modified Component in Soybeans

• Protocol of the Polymerase Chain Reaction for Detecting Genetically Modified Components in Potatoes

• Protocol of the Polymerase Chain Reaction for Detecting Genetically Modified Components in cotton

• Protocol of the real-time PCR for detecting genetically modified plants and their derived products

• Protocol of PCR for detection of genetically modified feed

• Protocol of the Polymerase Chain Reaction for Detecting Genetically Modified Components in maize

• Methods of sampling and preparation of samples for detection of genetically modified components in plants and their derived products

• Protocol of the Detection Methods for Genetically Modified tobacco

• Protocol of the polymerase chain reaction for detecting genetically modified in rapeseeds

• Protocol of the Qualitative Polymerase Chain Reaction for Detecting Genetically Modified Plant Components in Food

• Protocol of the Qualitative Polymerase Chain Reaction for Detecting Genetically Modified Plant Components in Edible Oil

8 GB standard protocols

• Detection of genetically modified organisms and derived products - Nucleic acid extraction

• Detection of genetically modified organisms and derived products – Gene chip detection

• Detection of genetically modified organisms and derived products – Protein based methods

•Detection of genetically modified organisms and derived products - Methods for sampling and sample preparation

• Detection of genetically modified organisms and derived products - Quantitative nucleic acid based methods

• Detection of genetically modified organisms and derived products - Qualitative nucleic acid based methods

•Detection of genetically modified organisms and derived products - General requirements for laboratories

• Detection of genetically modified organisms and derived products - General requirements and definitions

Construction of labs for GMOs detection

• 25 labs for GMOs detection

• Proficiency test

• Training of detection technology

Detection methods Information

National Genetically Modified Product Detection Laboratories and Detection Method Complex

Labs for GMOs detection

Construction of labs Requirements

for detection labs

Proficiency test of detection labs

Exchanging and training of detection technology

Methods for GMOs detection

DNA detection methods

Protein detection methods

Issue of standard protocols for GMOs

detection

Kits for GMOs detection

GMO websites

GMO databases

Thank you!