Unlock instant, AI-driven research and patent intelligence for your innovation.

Cold-resistance gene SiCOR3708 of herba saussureae involucratae and application thereof

A technology of gene and snow lotus, applied in application, genetic engineering, plant genetic improvement, etc., can solve the problems of laborious, time-consuming, lack of pertinence, etc., and achieve the effect of improving cold resistance

Inactive Publication Date: 2012-07-25
NANJING TECH UNIV +2
View PDF0 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there are three basic ways to solve cold damage: first, to adopt more advanced cultivation techniques; second, to improve existing plant varieties, especially those with high economic value, but traditional improvement methods are time-consuming, laborious, and Lack of pertinence; third, with the research of plant cold resistance mechanism, the cloning of cold resistance related genes and the development of transgenic technology, the research of transgenic plants with related recipient plants has made genetic engineering to cultivate high-efficiency cold-resistant varieties become an important Therefore, it is urgent to develop genes and proteins related to cold resistance

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
  • Cold-resistance gene SiCOR3708 of herba saussureae involucratae and application thereof
  • Cold-resistance gene SiCOR3708 of herba saussureae involucratae and application thereof
  • Cold-resistance gene SiCOR3708 of herba saussureae involucratae and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] Example 1: Cloning of the SiCOR3708 gene of Saussurea tianshanensis

[0018] The seedlings of Snow Lotus were cultivated at 4°C for 0.5 hours, the total RNA was extracted using Trizol reagent (Invitrogen Company), and the cDNA suppression subtraction kit (PCR-Select cDNA Subtraction kit, Clontech Company) was used to construct the same method as that of Snow Lotus in Tianshan according to its operation steps. For the cDNA subtraction library related to cold resistance, 1800 cDNAs were selected from them, and hybridized with the forward and reverse subtraction cDNAs respectively, and the cDNAs with forward hybridization signals but no reverse hybridization signals were selected for nucleotide sequence determination , the Saussurea sativa SiCOR3708 gene was obtained, the sequence is shown in Sequence Table 1, the gene is 632bp in full length, has a complete reading frame (28-588bp) and 5' untranslated region (1-27bp) and 3' untranslated region (589-632bp ), the reading fr...

Embodiment 2

[0019] Example 2: Construction of SiCOR3708 gene constitutive high-efficiency expression vector

[0020] The construction process of the expression vector pCHF3-SiCOR3708 is as follows: firstly, the full-length cDNA of the SiCOR3708 gene is obtained by amplifying the cDNA of Snow Lotus as a template. The amplification primers are as follows, with SmaI restriction sites at both ends:

[0021] Forward primer: 5'-CCCGGGATGAGTTATTCATACGATGA-3'

[0022] Reverse primer: 5'-GGGCCCCTTACTCATATCCCGCCTCCA-3'

[0023] The reaction conditions are: 95°C, 4min, 1 cycle; 94°C, 30sec, 54°C, 30sec, 72°C, 1min, 30 cycles; 72°C, 10min, end the reaction. After purifying the obtained amplified product, carry out SmalI enzyme digestion and purification, and carry out SmalI enzyme digestion, dephosphorylation reaction and purification on the carrier pCHF3 at the same time, and then perform ligation reaction, and first use restriction endonuclease to obtain the recombinant vector Perform double enzy...

Embodiment 3

[0027] Embodiment 3: Transformation of Agrobacterium with pCHF3-SiCOR3708 expression vector

[0028] Inoculate a single colony of Agrobacterium GV3101 in 50ml YEB medium, culture at 28°C, 200rpm shaking until OD 600 0.4, ice-bathed for 30 minutes, centrifuged at 4000rpm for 10 minutes, collected the bacteria and washed with 10ml 0.15M CaCl 2 suspended. Centrifuge again to collect the bacteria, suspend in 1ml 20mM CaCl 2 In the solution, take 100ul of the bacterial solution and put it into a sterile 1.5ml centrifuge tube, add 0.5ug of pCHF3-SiCOR3708 plasmid, mix gently, put in ice bath for 30 minutes, quickly freeze in liquid nitrogen for 1 minute, after melting at 37°C, add 1ml of YEB culture medium, cultivated at 28°C and 100rpm for 2 hours, centrifuged at 4000rpm for 30 seconds, discarded the supernatant, added 100ul of YEB culture medium, and spread the bacteria on the YEB solid medium containing 50ug / ml spectinomycin after 28 Cultivate until a single colony grows, and ...

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 cold-resistance gene SiCOR3708 of herba saussureae involucratae and application thereof for plants to resist cold. After herba saussureae involucratae tissue culture seedling is cultured for 0.5 hour at 4 DEG C, mRNA thereof is extracted, and the mRNA of herba saussureae involucratae aseptic seedling growing at 20 DEG C is extracted; a suppression substractive hybridization technology is adopted to obtain a subtracted library of herba saussureae involucratae low-temperature-induced cDNA, the sequence of the cDNA fragment in the subtracted library is determined to obtain the herba saussureae involucratae SiCOR3708 gene. The herba saussureae involucratae SiCOR3708 gene is composed of 632 basic groups, the coder frame of the gene is the basic groups from 5'end 28 to585, and a built SiCOR3708 gene expression carrier is transformed into arabidopsis to obviously improve the cold resistance capability of plants. The obtained cold-resistance protein gene of the herba saussureae involucratae is connected with the plant expression carrier and is induced into crops, such as cotton, wheat, rice, corn and the like so as to improve the cold-resistance performance of the plant, and thus the cold-resistance gene SiCOR3708 has wide application field.

Description

technical field [0001] The invention relates to the technical field of gene cloning and organism transformation in the field of biotechnology, specifically, the invention relates to the technical field of a cold-resistant gene of Saussurea chinensis and its application in plant cold resistance. Background technique [0002] Temperature is one of the important conditions for the normal growth and survival of plants, and it is also one of the important factors that limit the shape and distribution of plants. Low temperature damage to plants has become a global public hazard. my country has a large latitude span and diverse climates, and cold damage occurs from time to time. In recent years, cold damage has become one of the important natural disasters in my country in non-cold seasons, and it is gradually intensifying. It may become the largest natural disaster in agricultural production. At present, there are three basic ways to solve cold damage: first, to adopt more advanc...

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
Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29A01H5/00
Inventor 王志方艾秀莲李晨华徐虹欧阳平凯王博
Owner NANJING TECH UNIV