Herbicide tolerant, insect resistant maize

The maize has been produced through cross breeding of modified parental lines to confer tolerance to herbicides and resistance to insects. For Coleoptera resistance, the maize expresses Bacillus thuringiensis Cry3Bb1, which forms pores in the insect's midgut lining, leading to cell lysis and septicemia. Additionally, the maize contains an RNA interference cassette targeting Diabrotica virgifera virgifera (Western corn rootworm) DvSnf7, an essential cellular component of endosomal sorting complex required for transport. Instead of producing a protein, the cassette produces RNA molecules that cause gene silencing of this gene in western corn rootworm upon consumption of plant tissue and leads to mortality. For tolerance to glyphosate, the maize expresses Agrobacterium tumefaciens 5-enolpyruvylshikimate-3-phosphate synthase, a bacterial variant of an endogenous gene, which prevents the herbicide interference of the shikimate pathway (responsible for essential aromatic amino acid biosynthesis). Both parental lines contain a 5-enolpyruvylshikimate-3-phosphate synthase gene cassette. However, due to the general (non-specific) expression of one parental line, the tissue-specific expression is lost in the resulting cross.

PV-ZMAP1043; PV-ZMIR10871

DNA insert from MON87427 (MON-87427-7) vector PV-ZMAP1043
Transcription of Agrobacterium tumefaciens 5-enolpyruvylshikimate-3-phosphate synthase (epsps) commences from the Cauliflower mosaic virus (CaMV) enhanced 35S promoter and terminates at the A. tumefaciens nopaline synthase (nos) terminator. The transcript contains a Zea mays heat shock protein 70 (hsp70) intron, an Arabidopsis thaliana N-terminal chloroplast transit peptide sequence for chloroplast targeting of the protein and epsps. The CaMV enhanced 35S promoter-hsp70 combination promotes gene expression in female and vegetative tissues, but not in male reproductive tissues (pollen microspores and tapetum).

Note:
- Southern blot analyses indicate that a single copy of the T-DNA was inserted at a single site in the parental maize genome and no plasmid vector backbone sequences were detected to have been integrated. DNA sequencing analyses further indicated that the expected T-DNA sequences were integrated.
-The epsps coding sequence is the codon optimized coding sequence of the aroA gene from Agrobacterium sp. strain CP4 encoding EPSPS.

DNA insert from MON87411 (MON-87411-9) vector PV-ZMIR10871
The MON87411 genome contains an RNA interference (RNAi) cassette targeting Diabrotica virgifera virgifera, a Bacillus thuringiensis Cry3Bb1 cassette and an Agrobacterium tumefaciens 5-enolpyruvylshikimate-3-phosphate synthase (epsps) cassette.

Transcription of the RNAi cassette commences from the Cauliflower mosaic virus 35S enhanced promoter and terminates at the Pisum sativum ribulose bisphosphate carboxylase small chain 2 terminator. The transcript initially contains a Zea mays heat shock protein 70 intron, which contributes to enhanced expression in vegetative tissues of the plant, and two partial coding sequences of the D. virgifera virgifera Snf7p gene, which encodes the SNF7 subunit of the ESCRT-III complex. The two Snf7p sequences are in an inverted orientation, separated by a 150 nucleotide intervening sequence, which allows base pairing between the inverted sequences and hairpin RNA formation post-transcription, which then triggers an RNAi response. Due to RNAi processing, small interfering RNA molecules (roughly 21-23 nucleotides in length) will be produced and thus no translation into protein will occur from this cassette.

Transcription of the cry3Bb1 is under control of the Z. mays physical impedance induced protein promoter and Triticum aestivum (wheat) heat shock protein 17.3 terminator. The transcript also contains a wheat 5' untranslated leader from chlorophyll a/b-binding protein and Oryza sativa actin 1 intron for enhanced expression of the transgene. Expression of epsps is under control of an O. sativa alpha tubulin promoter and terminator. The transcript additionally contains Arabidopsis thaliana chloroplast targeting peptide 2 to sequester the protein to the chloroplast.

Note:
- Sequencing, PCR and bioinformatic analyses indicate that a single, intact insertions of the three gene cassettes occurred in the parental line.
- No plasmid backbone was detected.

Kindly refer to the parental LMO records for more information.