Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Method for improving resistance of rice to HPPD inhibitor herbicides through gene editing, and special sgRNA used therein

A technology of gene editing and transgenic rice, which is applied in the biological field, achieves the effect of obvious tolerance and significant application and promotion value

Active Publication Date: 2021-08-17
JIANGSU LIXIAHE REGION AGRI RES INST
View PDF2 Cites 3 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are still few studies on rice's own OsHPPD gene, and there is no report that resistance can be produced after changing a known site.

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Method for improving resistance of rice to HPPD inhibitor herbicides through gene editing, and special sgRNA used therein
  • Method for improving resistance of rice to HPPD inhibitor herbicides through gene editing, and special sgRNA used therein
  • Method for improving resistance of rice to HPPD inhibitor herbicides through gene editing, and special sgRNA used therein

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Example 1 Gene editing vector construction

[0027]According to the OsHPPD genome sequence, combined with Huazhong Agricultural University CRISPR-P 2.0 (http: / / crispr.hzau.edu.cn / CRISPR2 / ) to design targets and screen out 5 specific sequences with high predicted targeting efficiency and low off-target efficiency. for the generation of sgRNA. The target 1 sequence (SEQ ID NO.6) is located at 309-328 bp of the full-length 451 bp of the second exon of the OsHPPD gene gene (the nucleotide sequence is shown in SEQ ID NO.4). The PAM sequence TGG is located at the 3' end of the target sequence. Other sgRNA target sequences are listed in Table 1. Entrusted Wuhan Boyuan Biotechnology Co., Ltd. to use the CRISPR / Cas9 gene editing system to edit the target gene. The specific operation method refers to the literature Ma et al. Cas9 recombinant vector ( figure 1 ), the recombinant vector contains both the coding gene encoding the specific sgRNA and the coding gene of the Cas9 pr...

Embodiment 2

[0028] Example 2 Obtaining and Identification of OsHPPD Gene Edited Transgenic Plants

[0029] The constructed knockout vector pYLCRISPR / Cas9-OsHPPD plasmid was transformed into the japonica rice recipient variety Yangjing 3012 by using the genetic transformation method mediated by Agrobacterium EHA105. The genetic transformation of rice was entrusted to Wuhan Boyuan Biotechnology Co., Ltd. The general process of transformation is as follows: the pYLCRISPR / Cas9-OsHPPD plasmid was extracted and transformed into Agrobacterium EHA105, and the Agrobacterium EHA105 containing the pYLCRISPR / Cas9-OsHPPD plasmid was used to infect the callus of the japonica rice recipient variety Yangjing 3012, and then transferred to Cultured in the dark at 26°C for 3 days on the co-culture medium, the washed callus was transferred to the selection medium containing hygromycin for resistance selection, and the selected resistant callus was transferred to the pre-differentiation medium for 14 days. T...

Embodiment 3

[0035] Example 3 OsHPPD Gene Editing Transgenic Lines Identification of Herbicide Mesotrione Resistance

[0036] The 2 homozygous gene editing Ts created by target 1 in Example 2 0 Generation plants and other transgenic plants were added to T 2 Homozygous strains were obtained. Will T 2 Rinse the seeds of generation strains and wild-type rice with clean water several times, and use the method of "soaking three times and accelerating one" to soak the seeds for germination acceleration, that is, soak the seeds for three days and accelerate germination for one night. , pay attention to heat preservation during the germination period, and pay attention to temperature control after emergence to prevent seedlings from jumping high. When the seedlings grow to the 2-3 leaf stage, spray the mesotrione suspension. The mesotrione suspension contains 10 μmol / L mesotrione Ketone and 0.02% Tween-20, spray until the blade surface is all wet. After 12 days, check the phenotype of the rele...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses a method for improving resistance of rice to HPPD inhibitor herbicides through gene editing and a special sgRNA used therein. The coding sequence of the specific sgRNA for editing a rice OsHPPD gene based on CRISPR / Cas9 in the invention is as shown in SEQ ID NO. 6. A CRISPR / Cas9 vector aiming at the rice OsHPPD gene contains a coding gene sequence for coding the specific sgRNA. A method for improving the resistance of rice to the HPPD inhibitor herbicides through gene editing comprises the following step: introducing the coding gene of the specific sgRNA and the coding gene of a Cas9 protein into starting rice to obtain transgenic rice with a mutated OsHPPD gene. Compared with the starting rice, the resistance of the transgenic rice to the HPPD inhibitor herbicides is improved.

Description

technical field [0001] The invention belongs to the field of biotechnology, and specifically relates to a method for improving rice resistance to HPPD inhibitor herbicides through gene editing and a special sgRNA thereof. Background technique [0002] Rice is one of the main food crops in my country, and weed damage is one of the important factors restricting rice production. There are as many as 40 kinds of common and most common weeds in rice fields in my country, and the area of ​​weed damage in rice fields is 3.8 million ha per year 2 , Accounting for more than 11.5% of the rice planting area, the annual loss of rice due to grass damage is as high as more than 10 million tons, and the damage of weeds has seriously affected the yield and quality of rice. [0003] At present, in addition to traditional manual weeding, the most important field weeding method is chemical control, which has the advantages of high efficiency, economy, and labor saving, and plays an important ...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): C12N15/113C12N15/84C12N15/55C12N15/53A01H5/00A01H6/46
CPCC12N15/8218C12N15/8274C12N15/8205C12N9/22C12N9/0069C12Y113/11027
Inventor 吴云雨李爱宏肖宁潘存红蔡跃余玲李育红张小祥刘广青黄年生周长海戴正元季红娟刘建菊陈梓春时微王志平
Owner JIANGSU LIXIAHE REGION AGRI RES INST
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com