Analysis of Primary Biotechnology Literature

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Analysis of Primary Biotechnology Literature
Biotechnology Education Forum NDSU Extension
Inservice Training Bismarck, ND March 1, 2002
Phil McClean Department of Plant Science North
Dakota State University
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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 source
• Does the pollen affect the butterfly
• Losey et al. (1999) Nature 399214
• Yes!!
• Sears et al. (2001) PNAS 9811937
• No!!

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Concerns for Biotech Products Inadvertent Gene
Transfer

• Wild relatives receive transgene from biotech
  crops
• Important genetic diversity will be lost
• Has it been observed?
• Quist Chapela (2001) Nature 414541
• Yes!!
• Has it been challenged?
• Christou (2002) Transgenic Research 11iii
• Yes!!

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Lets Review the Primary Literature
Transgenic DNA Introgressed Into
Traditional Maize Landraces in Oaxaca, Mexico
David Quist and Ignacio Chapela
Nature 414541-543 November 29, 2001
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What does the title tell us?
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What does the title tell us?
Transgenes were found in landraces
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What does the title tell us?
Transgenes were found in landraces
What does this imply?
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What does the title tell us?
Transgenes were found in landraces
What does this imply?
Biotech crops have contaminated nature
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Paragraph 1 What was analyzed?
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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)

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Paragraph 1 How were samples analyzed?
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Paragraph 1 How were samples analyzed?

• DNA isolated from flour
• Scored for presence of 35S promoter
• Why???
• Widely used transgene contruct element
• Polymerase chain reaction technique
• Presence of product
• transgene promoter DNA is in sample

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Paragraph 2 What are the results?
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Paragraph 2 What are the results?

• Amplification observed in 5/7 samples
• Weak, but present
• Low copy gave low signal
• Store sample gave strong signal
• Bt1 and RR1 gave strong signal
• Historical and Peruvian samples negative
• Positive control amplification observed in all
  samples

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Paragraph 2 Supporting evidence?
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Paragraph 2 Supporting evidence?

• Independent confirmation by Mexican govt
• Oaxaca and one other state
• Samples near authors site also positive at low
  levels
• Mexican experiment analyzed individual kernels
• Strong signal than pooled samples authors used

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Paragraph 3 Confirming Experiment?
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Paragraph 3 Confirming Experiment?

• Weak signal amplified with internal primers
• New fragment sequenced
• Sequence equal Monsanto 35S promoter sequence

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Paragraph 4 Other genes present?
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Paragraph 4 Other genes present?

• Samples asssayed for other transgenes
• NOS terminator sequence
• A3, B2 and K1
• Bt toxin gene (cryIAb)
• B3
• Conclusion
• Multiple transgenes found in the Mexican
  landraces

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Paragraph 5 Clues to transgene origin?
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Paragraph 5 Clues to transgene origin?

• Samples assayed for genomic location of transgene
• Procedure Inverse PCR
• Scores transgene and neighboring maize DNA
• Four samples contain known flanking genes
• A2, A3, B3, K1
• Transgene located in same position as Novartis
  Bt11
• Samples A3, K1
• Conclusion
• GMOs to landraces introgression occurring
• Despite GMO planting in Mexico (1998-now)

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The Challenge to Quist Chapela

• Chistou (2002)
• Fundamental flaws (experimental design) in
  research
• Results can be explained by
• Contaminated samples
• Flawed iPCR interpretation
• Cross pollination not supported by results
• Better experiment
• Grow plant samples out and
• Score phenotype (Bt or Glyphosate tolerant)
• Score genotype (screening individual plants)

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Whats Next??

• Funding becomes available
• Detailed experiments
• New results or interpretation
• Whats at stake?
• Integrity of biotech approaches to crop
  improvement