DNA insert from PV-ZMAP1043
Transcription of Agrobacterium tumefaciens
5-enolpyruvylshikimate-3-phosphate synthase (cp4 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 cp4 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).
- 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 cp4 epsps coding sequence is the codon optimized coding
sequence of the aroA gene from Agrobacterium sp.
strain CP4 encoding CP4 EPSPS.
DNA insert from PV-ZMIR245
Two insecticidal protein expression cassettes were inserted into
the genome. Bacillus thuringiensis cry1A.105 expression is
under the control of the CaMV 35S enhanced promoter, which first
transcribes wheat (Triticum aestivum) 5' untranslated
region of the chlorophyll a/b-binding protein (cab) and a
rice actin 1 intron before transcribing cry1A.105.
Transcription terminates at the wheat heat shock protein 17.3
terminator. Expression of the B. thuringiensis cry2Ab2
starts at the Figwort mosaic virus (FMV) promoter, which
transcribes the Zea mays heat shock protein 70
(hsp70), then the Z. mays transit peptide and the
cry2Ab2 coding sequence, before terminating at the
- The Cry2Ab2 coding sequence was modified for optimal expression
- South blot analysis confirmed that single insertions of both
cry2Ab2 and cry1A.105, as well as no vector
backbone were present and in the parent.
- A deletion removed the duplicated enhancer elements compared to
the original CaMV e35S promoter in PV-ZMIR245.
- The selectable marker, nphII, cassette was bred out of
the parental line and became not associated with this
DNA insert from PV-ZMHT507801
The Streptomyces viridochromogenes phosphinothricin
N-acetyltransferase (pat) gene is under the control of the
Andropogon gerardii ubiquitin promoter and the
Oryza sativa alpha-amylase/trypsin inhibitor terminator.
The transcript includes the A. gerardii 5' untranslated
leader sequence and an intron from ubiquitin before (5') coding
sequence of pat.
The Stenotrophomonas maltophilia dicamba monooxygenase
(dmo) gene is under control of the Peanut chlorotic
streak caulimovirus (PC1SV) full-length transcript promoter
and the Triticum aestivum (wheat) heat shock protein 17
terminator. The transcript produced contains a wheat chlorophyll
a/b-binding 5' untranslated leader sequence (for improved gene
expression), an O. sativa actin 1 untranslated region and
intron (for improved gene expression), the untranslated and
targeting region of Petunia hybrida chloroplast transit
peptide 4 (for chloroplast targeting of the protein) and
- Originally, the plasmid vector contained two T-DNA elements that
were inserted during the initial transformation event: one
containing the dmo and pat expression cassettes,
and a second containing an Agrobacterium tumefaciens
5-enolpyruvylshikimate-3-phosphate synthase (cp4-epsps)
expression cassette. The cp4-epsps expression cassette is
regulated by the O sativa actin 1 promoter and 5′
untranslated leader, O. sativa intron, the Arabidopsis
thaliana chloroplast targeting peptide 2 targeting sequence,
and the A. tumefaciens nopaline synthase 3′
untranslated region. Subsequent traditional breeding, segregation,
selection, and screening were used to isolate those plants that
contain the dmo and pat expression cassettes
(T-DNA I) and do not contain the cp4-epsps expression
cassette (T-DNA II).
- Molecular characterization of MON87419 indicated that a single
copy of T-DNA I was integrated into the maize genome at a single
intact locus that includes all expected elements within the insert,
with the exception of incomplete Right and Left Border sequences.
These analyses also showed no PV-ZMHT507801 backbone elements or
T-DNA II sequences were present in the event.
DNA insert from PV-ZMGT32
The plant expression plasmid vector, PV-ZMGT32 contains two
adjacent plant gene expression cassettes each containing a single
copy of the cp4 epsps. In the first (5' end) expression
cassette, the cp4 epsps gene is under the transcriptional
regulation of an Oryza sativa actin promoter and a
nos terminator. An O. sativa actin intron is also
present in the transcript for enhanced expression of the coding
sequence. The second cassette consists of another cp4
epsps gene regulated by an CaMV enhanced 35S promoter
(containing a duplicated enhancer region) and a nos
terminator. Similarly, an intron from the maize heat shock protein
70 (hsp70) was included for enhancing expression of the
coding sequence. Both promoters of the gene cassettes are expected
to promoter high levels of transcription.
- The parental NK603 line contained a single, intact insertion
containing both cp4 epsps gene cassettes.
- Due to restriction digest prior to particle bombardment, the
vector backbone, containing E. coli neomycin
phosphotransferase II and origin of replication, were not
incorporated into the parental genome.
For more information, kindly refer to the parental line