Methods and results
Detection of transgenic DNA
The authors looked for three specific fragments of transgenic DNA
in their PCR analyses:
- the Cauliflower Mosaic Virus 35S promoter (CaMV 35S), which
acts as an on-off switch for transgenes in most GE varieties.
- the cryIAb gene, which encodes an insecticidal protein
found in many Bt varieties.
- the nos termination sequence used to mark the end of
(See How Do You Make a Transgenic
Plant? for details of transgene construction).
The CaMV 35S sequence was detected in four of the six landrace
samples, although with a weak intensity compared to the sample from
the government food agency and the known GE samples. The authors
also reported detecting the nos sequence in two of six landrace
samples, and the cryIAb sequence in one landrace sample.
As expected, these sequences were not detected in the GE-free samples.
The weak intensity of CaMV 35S amplification in the landraces indicated,
according to the authors, that the transgenic DNA was present in
a small proportion of kernels. This is consistent with similar analyses
conducted by the Mexican government, which found that 3 to 10% of
kernels from the same area contained transgenic DNA (Quist
and Chapela, 2001). To confirm the identity of the weak CaMV
35S DNA bands amplified from the landraces, they were subjected
to another round of PCR amplification to increase the quantity of
DNA, then analyzed to obtain their nucleotide sequences. In all
cases, the sequence obtained from the landrace DNA bands matched
the sequence of the suspected CaMV 35S promoter.
Evidence for transgene movement in the genome
Quist and Chapela performed inverse PCR (iPCR) to examine the regions
flanking the CaMV 35S DNA, and thus to determine where in the genome
the transgenic DNA had integrated. The iPCR procedure allows amplification
of DNA segments that are adjacent to DNA of known sequence (http://www.pmci.unimelb.edu.au/core_facilities/manual/mb390.asp;
et al., 1988). Sequencing of iPCR products yielded several different
results, which the authors claimed indicated insertion of the transgenic
DNA into numerous sites in the genome. The authors stated that two
of the sequences allegedly flanking CaMV 35S had similarity to part
of the adh1 gene used in several Bt maize lines to enhance
gene expression, and other sequences were similar to non-transgenic
maize DNA. This led Chapela and Quist to conclude that the transgenic
construct was capable of inserting into the genome and then moving
around either during the original transformation event or during
the genetic recombination that occurs each time a seed is produced.
This assertion is significant because if the CaMV 35S promoter were
to wander through the genome it could turn on genes that are not
normally active, and disrupt, and therefore turn off, genes that
are necessary for normal growth and development.