Purpose of insecticidal protein
A protein and pest technology, applied in the field of insecticidal protein
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no. 1 example
[0082] The first embodiment, the acquisition and synthesis of genes
[0083] 1. Obtain the nucleotide sequence
[0084] The amino acid sequence (789 amino acids) of Vip3A-01 insecticidal protein, as shown in SEQ ID NO:1 in the sequence listing; The Vip3A-01 nucleotide sequence ( 2370 nucleotides), as shown in SEQ ID NO:2 in the sequence listing. Amino acid sequence (789 amino acids) of Vip3A-02 insecticidal protein, as shown in SEQ ID NO:3 in the sequence listing; Vip3A-02 nucleotide sequence ( 2370 nucleotides), as shown in SEQ ID NO:4 in the sequence listing.
[0085] The amino acid sequence (615 amino acids) of Cry1Ab insecticidal protein, as shown in SEQ ID NO:7 in the sequence listing; Cry1Ab nucleotide sequence (1848 nucleotides) of encoding corresponding to the amino acid sequence of said Cry1Ab insecticidal protein , as shown in SEQ ID NO:8 in the sequence listing.
[0086] The amino acid sequence (1436 amino acids) of Cry1Ab+Vip3A insecticidal protein, as shown in...
no. 2 example
[0089] The second embodiment, construction of recombinant expression vector and transformation of recombinant expression vector into Agrobacterium
[0090] 1. Construction of a recombinant cloning vector containing the Vip3A gene
[0091] The synthetic Vip3A-01 nucleotide sequence was connected to the cloning vector pGEM-T (Promega, Madison, USA, CAT: A3600), and the operation steps were carried out according to the instructions of the pGEM-T vector produced by Promega Company to obtain the recombinant cloning vector DBN01-T , its construction process is as follows figure 1 Shown (wherein, Amp represents the ampicillin resistance gene; f1 represents the replication origin of phage f1; LacZ is the LacZ initiation codon; SP6 is the SP6 RNA polymerase promoter; T7 is the T7 RNA polymerase promoter; Vip3A-01 is the Vip3A -01 nucleotide sequence (SEQ ID NO: 2); MCS is the multiple cloning site).
[0092]Then, the recombinant cloning vector DBN01-T was transformed into Escherichia...
no. 3 example
[0105] The third embodiment, the acquisition of transgenic plants
[0106] 1. Obtaining transgenic corn plants
[0107] According to the routinely used Agrobacterium infection method, the calli of the aseptically cultured corn variety Zong 31 (Z31) was co-cultured with the Agrobacterium described in 3 in the second embodiment to construct 2 in the second embodiment T-DNA in the recombinant expression vectors DBN100002 and DBN100003, DBN100740 and DBN100736 (comprising the promoter sequence of corn Ubiquitin gene, Vip3A-01 nucleotide sequence, Vip3A-02 nucleotide sequence, Cry1Ab+Vip3A nucleotide sequence, The Cry1Ab nucleotide sequence, the Hpt gene and the Nos terminator sequence) were transferred into the corn genome, and the corn plants transferred to the Vip3A-01 nucleotide sequence, the corn plant transferred to the Vip3A-02 nucleotide sequence, and the transferred Maize plants with Vip3A-01-Cry1Ab nucleotide sequence and maize plants transformed with Cry1Ab+Vip3A nucleo...
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