Citrus canker susceptible gene CsLOB

A technology for citrus canker and susceptible genes, applied in the field of citrus canker susceptible gene CsLOB, can solve the problem of not finding a susceptible gene, and achieve the effect of great application value

Active Publication Date: 2014-06-25
SHANGHAI JIAO TONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In citrus canker, previous studies fa

Method used

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  • Citrus canker susceptible gene CsLOB
  • Citrus canker susceptible gene CsLOB
  • Citrus canker susceptible gene CsLOB

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0018] Cloning of CsLOB gene promoter

[0019] In this example, the 500 bp DNA fragment upstream of the CsLOB gene in citrus and the PthA gene in X. citri were cloned, and the interaction with the PthA protein of X. citri was verified by the transient expression technology of Agrobacterium. Using citrus genomic DNA as a template, primers pCsLOB-F (SEQ ID NO.4): 5'-AAAGGTACCATGACGGTGGGAGCCGGGGTG-3' and pCsLOB-R (SEQ ID NO.5): 5'-TCTAGAGCCTGGTGATACTTCCTTGCCAGAA-3' were designed for PCR amplification. Increase, the PCR amplification conditions are: 95°C / 5min+(94°C / 45sec+55°C / 30sec+72°C / 1min30sec)×38 cycles+72°C / 10min. After the PCR product was digested by SalI and EcoRI, it was connected to the pCAMBIA1381 vector to obtain pCAMBIA1381-pCsLOB. With the help of Agrobacterium EH105, it was constructed with the PHB vector expressing PthA on tobacco for transient expression at the same time, and it took 2-3 days for the transformation of tobacco After culturing in a greenhouse (25°C,...

Embodiment 2

[0021] Pathogenicity of CsLOB gene to citrus after specific activation

[0022] In this example, the expression of the CsLOB gene is activated by artificially synthesizing the tal gene. A segment of the sequence (SEQ ID NO.6) of the selected CsLOB promoter was designed and synthesized as a target point dtal gene (see figure 1 sequence in blue). Using the PthA gene sequence of X. citri as a template, primers dTAL-F (SEQ ID NO.7): 5'-GGCGGTCGACCGGCGCTGGAGAGCAT-3' and dTAL-R (SEQ ID NO.8): 5'-AATGCATGCAAAGACGCCTGGTCCG-3' were designed for For the PCR reaction, the amplification conditions are: 95°C / 5min+(94°C / 45sec+53°C / 45sec+72°C / 20sec)×32 cycles+72°C / 10min. The PCR product was recovered, and ligated to the pUFR034 vector together with the dtal gene fragment after being restricted by SalI and SphI to obtain the vector pUFR034-dTAL. The pUFR034-dTAL vector was transformed into X. citri without any tal using electroporation. Injection of citrus, X. citri containing the dtal ge...

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Abstract

The invention provides a citrus canker susceptible gene CsLOB. The sequence of the citrus canker susceptible gene CsLOB in citrus is as shown in SEQ ID No.1, the sequence length of a promoter is 500bp, and the sequence length of an ORF (Open Reading Frame) is 711bp; and the sequence of CsLOB protein is as shown in SEQ ID No.2. UPT box sequence which can be specifically combined with a citrus canker bacterium PthA exists in the CsLOB gene promoter, the sequence of the UPT box is as shown in SEQ ID No.3, and the length of the UPT box is 18pb. The obtained CsLOB gene can be specifically recognized by main toxic protein PthA derived from the citrus canker bacterium and is activated to be transcribed, so that the occurrence of citrus canker is caused. A new germplasm of citrus or a new material with durable resistance can be obtained through gene silencing of the CsLOB gene and genetic modification of the UPT box.

Description

technical field [0001] The invention relates to genes, in particular to a citrus canker susceptibility gene CsLOB. Background technique [0002] Citrus canker occurs widely in citrus growing areas in China, especially in Guangdong, Guangxi, Hunan and Fujian. When it is serious, it will cause leaf and fruit drop, poor plant growth, and affect yield and quality. [0003] X. citri can infect the host plant citrus and cause symptoms near the infection point. The mechanism is that certain genes of citrus are abnormally overexpressed, so that X. citri successfully obtains the required nutrients from the plant. During the recognition process of X. citri and host plants, X. citri secretes a variety of type III effector proteins into plant cells through the type III secretion system, among which PthA is considered to be the main virulence factor causing citrus canker. PthA belongs to TALE (Transcriptional Activator Like Effector) proteins, and its structure is conserved at the nitr...

Claims

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Application Information

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IPC IPC(8): C12N15/29C12N15/113C07K14/415
Inventor 陈功友李争邹丽芳
Owner SHANGHAI JIAO TONG UNIV
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