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Cotton plant event ac20-3 and primers and methods for its detection

A technology of cotton and cotton genome, applied in botany equipment and methods, biochemical equipment and methods, angiosperms/flowering plants, etc., can solve the problems of increased heat absorption and heat conductivity of transgenic cotton, and improve cotton fiber quality , Molecular characteristics are clear, the effect of genetic stability

Active Publication Date: 2020-01-31
BIOCENTURY TRANSGENE CHINA
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

John et al. (1996) introduced the acetyl CoA reductase gene (phaB) and the PHB synthase gene (phaC) into upland cotton cultivars. Although the fiber quality of transgenic cotton, such as strength and length, was not significantly improved, the absorption of transgenic cotton Significantly increased thermal and thermal conductivity

Method used

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  • Cotton plant event ac20-3 and primers and methods for its detection
  • Cotton plant event ac20-3 and primers and methods for its detection
  • Cotton plant event ac20-3 and primers and methods for its detection

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Example 1. Plant expression vector construction

[0030] Since the gene encoding the cellulose binding domain CBD (CBD) is derived from Clostridium cellulophorum, its GC content and codon preference are different from those of the cotton genome, so its base composition was changed according to the cotton codon preference, and artificially synthesized The method for synthesizing SP and CBD gene fusion sequence (sequence shown in SEQ ID No: 1), wherein the SP sequence is from the N-terminal 24 bases of Arabidopsis CEL1 gene, is the guide peptide sequence. The fusion sequence was entrusted to Takara Company to carry out the artificial synthesis of the whole gene, and the new gene was named SP+mCBD. The plant binary expression vector pCAMBIA2300 (purchased from Beijing Dingguo Changsheng Biotechnology Co., Ltd.) was selected as the plant expression vector, and the 35S promoter of the double enhancer of the NPTII gene was replaced with the Pnos promoter to reduce the express...

Embodiment 2

[0034] Example 2. Genetic transformation of upland cotton (Gossypium hirsutum):

[0035] Using the genetic transformation method mediated by Agrobacterium, the hypocotyl of Jimian 14 was transformed with the vector containing mCBD gene.

[0036] Agrobacterium LBA4404 containing the p2300-pE200-mCBD vector was picked and inoculated into kanamycin (kanamycin, km) 50 mg / L, rifampicin (rifampicin, rif) 50 mg / L and streptomycin (streptomycin, S / Sm) 50mg / L LB liquid culture medium, 28 ℃ shaking dark culture overnight to the logarithmic phase of bacterial growth. Dilute the bacterial solution with LB or YEB liquid medium at a ratio of 1:50 to 1:100 of bacterial solution:medium, then shake and culture for 4 to 6 hours, and dilute the bacterial solution to an OD600 value of 0.8 to 1.0.

[0037] The transgenic recipient material is Jimian 14. The hypocotyls of sterile seedlings grown for 3 to 4 days were taken, cut into 0.6 to 0.8 cm sections, dipped for 10 to 15 minutes, and the hypo...

Embodiment 3

[0038] Example 3. Screening and identification of T1 generation transgenic materials:

[0039] 215 individual plants of 41 lines of aC series cotton in the T1 generation were harvested, and the harvested cotton was tested for clothing and fiber, and the results were further statistically analyzed, and the transformation event aC20-3 was screened. Compared with the control, the micronaire value of the cotton fiber in the transformation event aC20-3 decreased by 23.2%, the specific strength increased by 12.1%, the cloth value increased by an average of 8.4%, and the length did not change significantly. The specific results are shown in Table 1.

[0040] Table 1

[0041]

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Abstract

The invention provides a cotton transformation event aC20‑3 for improving cotton fiber quality, its sequence, reagents and methods for its detection. Cotton plants carrying the aC20-3 event contain the event, the junction of the exogenous inserted DNA sequence and the cotton genomic DNA sequence, on the D5 chromosome. Utilizing the exogenous insert DNA sequence and the DNA sequence of the junction region on the flanking cotton genome, detection primers or probes can be designed for specific detection of aC20‑3 events. The detection method for the aC20-3 event can provide a convenient means for tracking the specific gene insertion event for the application of the plant event for breeding, and the specific gene insertion event can be used as a molecular marker to improve the efficiency of breeding work.

Description

technical field [0001] The invention belongs to the field of plant molecular biology, especially the field of transgenic crop breeding in agricultural biotechnology research, and particularly relates to a cotton transformation event aC20-3 with improved cotton fiber quality, and a unique method for detecting the transformation event. Background technique [0002] Cotton fiber is the main product of cotton production, and the output and quality of cotton fiber directly determine the output value and benefits of cotton production. Therefore, improving cotton fiber quality has always been the main goal of cotton breeding. However, the yield and quality of cotton are both quantitative traits, which are affected by many factors and there is a negative correlation between yield and quality, which seriously hinders the simultaneous improvement of cotton fiber quality and yield. Cotton cultivars are mainly upland cotton, and good fiber quality genes are mainly derived from diploid ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C12N15/63C12N1/21C12N15/11C12Q1/6895A01H5/00A01H6/60
CPCC12Q1/68C12N15/11A01H1/045
Inventor 王建胜何云蔚崔洪志
Owner BIOCENTURY TRANSGENE CHINA