Insect resistant, Drought tolerant maize (Water efficient maize for Africa)

The  modified maize was produced through the crossing of two modified parental lines (MON810 and MON87460) to achieve insect resistance and drought tolerance. For resistance to Lepidoptera pests, the maize expresses Bacillus thuringiensis Cry1Ab, which causes pore formation in the midgut epithelial cells of larvae and results in loss of gut lining integrity and eventual death. For drought tolerance, the maize expresses Bacillus subtilis cold shock protein, which binds RNA and maintains cellular functions under water-limited conditions (improvement of natural abiotic stress responses). The maize also contains an Escherichia coli neomycin phosphotransferase II cassette, which allowed for kanamycin selection during transformation of the MON87460 parental line.

PV-ZMBK07 and PV-ZMAP595

DNA insert from PV-ZMBK07
A partial insert containing Bacillus thuringiensis cry1Ab was inserted into the parental maize genome. Transcription is directed from the Cauliflower Mosaic Virus 35S enhanced promoter. The transcript contains a Zea mays heat shock protein 70 (ZmHsp70) intron and the coding sequence of cry1Ab. ZmHsp70 enhances expression of cry1Ab.

Note:
- The coding sequence of cry1Ab has been codon optimized for expression in plants. The codon optimization did not result in any changes to the amino acid sequence relative to the native sequence.
- Southern blot analysis indicated that a single partial insert is found within the parental genome.
- Southern blot analysis did not detect the presence of the Escherichia coli neomycin phosphotransferase II gene nor any DNA from plasmid PVZMGT10 (containing genes for glyphosate tolerance - cp4 epsps).
- ELISA protein analysis and feeding assays indicated expression of Cry1Ab.

DNA insert from PV-ZMAP595
The T-DNA insert contains the following gene cassettes: Bacillius subtillus cold shock protein (cspB) and Escherichia coli neomycin phosphotransferase II (nptII). 

Transcription of cspB is under control of the Oryza sativa actin 1 promoter and Agrobacterium tumefaciens trasncript 7 gene 3' untranslated region. The transcript initially contains an O. sativa actin 1 intron for enhanced gene expression of cspB. The sequence is removed (spliced) prior to protein translation. Constitutive expression of cspB is expected due to the actin promoter.

Transcription of nptII is under control of the Cauliflower Mosaic Virus (CaMV) 35S promoter and A. tumefaciens nopaline synthase terminator. High levels of transcription are expected due to the CaMV promoter.

Note:
- The coding seuquence of cspB has been codon optimized for optimal expression within plant cells.
- Southern blot analysis indicated that no vector backbone sequences were inserted into the parental genome
- Southern blot analysis indicated that the parental genome contains a single insertion
- Sequencing analyses confirm the Southern blot analyses.
- A 22 base pair deletion of genomic DNA at the insert-to-plant DNA junction occurred.
- loxP sites can be found in the parental genome and could potentially allow for the excision of the nptII cassette by CRE recombinase.

For more information, kindly refer to the parental records.

The Water Efficient Maize for Africa (WEMA) project is a public-private partnership led by the African Agricultural Technology Foundation (AATF) to address the effects of drought on farmers. The project partners have been developing new drought-tolerant African maize varieties since 2008, incorporating conventional advanced plant breeding and biotechnology.