NDSU

Extension

Analysis of Primary Biotechnology Literature

Phil McCleanDepartment of Plant Science

North Dakota State University

Biotechnology Education ForumNDSU Extension Inservice Training

Bismarck, NDMarch 1, 2002

NDSU

Extension

Concerns for Biotech Products:Ecological Balance Will Be Altered

Other organisms will be affected

•Monarch butterfly

•Maize Bt pollen falls on neighboring milkweed

•Milkweed a monarch butterfly food sourceDoes the pollen affect the butterfly

•Losey et al. (1999) Nature 399:214

Yes!!

•Sears et al. (2001) PNAS 98:11937

No!!

NDSU

Extension

Concerns for Biotech Products:Inadvertent Gene Transfer

Wild relatives receive transgene from biotech cropsImportant genetic diversity will be lostHas it been observed?

•Quist & Chapela (2001) Nature 414:541

Yes!!Has it been challenged?

•Christou (2002) Transgenic Research 11:iii

Yes!!

NDSU

Extension

Transgenic DNA Introgressed Into TraditionalMaize Landraces in Oaxaca, Mexico

David Quist and Ignacio Chapela

Nature414:541-543

November 29, 2001

Let’s Review the Primary Literature

NDSU

Extension

What does the title tell us?

NDSU

Extension

What does the title tell us?

Transgenes were found in landraces

NDSU

Extension

What does the title tell us?

Transgenes were found in landraces

What does this imply?

NDSU

Extension

What does the title tell us?

Transgenes were found in landraces

What does this imply?

Biotech crops have contaminated nature

NDSU

Extension

Paragraph 1: What was analyzed?

NDSU

Extension

Paragraph 1: What was analyzed?

Bulk seed from corn cobs from landraces

•Fields isolated from roads

•Six samples (A1-A3 and B1-B3)Store sample (K1)Controls

•Peruvian sample (P1)

•Oaxacan sample from 1971 (H1)

•Monsanto “Yieldgard” maize (Bt1)

•Monsanto “Roundup-Ready maize (RR1)

NDSU

Extension

Paragraph 1: How were samples analyzed?

NDSU

Extension

Paragraph 1: How were samples analyzed?

DNA isolated from flourScored for presence of 35S promoter

•Why???

•Widely used transgene contruct elementPolymerase chain reaction technique

•Presence of product = transgene promoter DNA is in sample

NDSU

Extension

Paragraph 2: What are the results?

NDSU

Extension

Paragraph 2: What are the results?

Amplification observed in 5/7 samples

•Weak, but present

•Low copy # gave low signalStore sample gave strong signalBt1 and RR1 gave strong signalHistorical and Peruvian samples negativePositive control amplification observed in all samples

NDSU

Extension

Paragraph 2: Supporting evidence?

NDSU

Extension

Paragraph 2: Supporting evidence?

Independent confirmation by Mexican gov’t

•Oaxaca and one other state

•Samples near authors site also positive at low levelsMexican experiment analyzed individual kernels

•Strong signal than pooled samples authors used

NDSU

Extension

Paragraph 3: Confirming Experiment?

NDSU

Extension

Paragraph 3: Confirming Experiment?

Weak signal amplified with internal primersNew fragment sequencedSequence equal Monsanto 35S promoter sequence

NDSU

Extension

Paragraph 4: Other genes present?

NDSU

Extension

Paragraph 4: Other genes present?

Samples asssayed for other transgenes

• NOS terminator sequence

A3, B2 and K1

• Bt toxin gene (cryIAb)

B3Conclusion:

Multiple transgenes found in the Mexican landraces

NDSU

Extension

Paragraph 5: Clues to transgene origin?

NDSU

Extension

Paragraph 5: Clues to transgene origin?

Samples assayed for genomic location of transgene

•Procedure: Inverse PCRScores transgene and neighboring maize DNAFour samples contain known flanking genes

•A2, A3, B3, K1Transgene located in same position as Novartis Bt11

•Samples A3, K1Conclusion:

GMOs to landraces introgression occurring

Despite GMO planting in Mexico (1998-now)

NDSU

Extension

The Challenge to Quist & Chapela

Chistou (2002)Fundamental flaws (experimental design) in researchResults can be explained by:

•Contaminated samples

•Flawed iPCR interpretationCross pollination not supported by resultsBetter experiment

•Grow plant samples out and:

Score phenotype (Bt or Glyphosate tolerant)

Score genotype (screening individual plants)

NDSU

Extension

What’s Next??

Funding becomes available•Detailed experiments•New results or interpretation

What’s at stake? Integrity of biotech approaches to crop improvement