Method for reducing the content of poplar lignin with RNA interference

An RNA interference and lignin technology is applied in the field of reducing the Klason lignin content of poplar trees, which can solve the problems of weak inhibition of endogenous gene expression, poor genetic stability of the inhibitory effect, and reduction of target gene expression, etc. Facilitate agronomic traits, high genetic stability effect

Inactive Publication Date: 2009-12-16
ANHUI AGRICULTURAL UNIVERSITY
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Problems solved by technology

However, the application of antisense technology has certain limitations. Its inhibition of endogenous gene expression is weak, and it often produces transitional phenotypes, which will hinder the accurate judgment of the function of the target gene.
The genetic stability of its inhibitory effect is poor, and it cannot stably and reliably reduce the expression of the target gene
[0006] 2. Gene knockout technology an

Method used

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  • Method for reducing the content of poplar lignin with RNA interference
  • Method for reducing the content of poplar lignin with RNA interference
  • Method for reducing the content of poplar lignin with RNA interference

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Experimental program
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Effect test

Embodiment Construction

[0022] 1. Obtaining CCR gene-specific RNAi fragments

[0023] 1.1 Extraction of Poplar Genomic DNA Using Improved CTAB Method

[0024] (1) Wash the leaves, remove the veins, weigh 1 g, add quartz sand, PVP (polyvinylpyrrolidone), and liquid nitrogen, grind them and put them into two 5mL Dorf tubes.

[0025] (2) Add 195 μL β-mercaptoethanol.

[0026] (3) Add 1950 μL of 1.5% CTAB (cetyltrimethylammonium bromide) and shake well.

[0027] (4) 65°C water bath for 20 minutes, gently shake once every 5 minutes.

[0028] (5) Add 1500 μL of chloroform / isoamyl alcohol (24:1) mixture, shake gently, and centrifuge at 10,000 rpm for 10 min.

[0029] (6) Take the supernatant, add 195 μL of 10% CTAB in a 60° C. water bath, 1950 μL of chloroform / isoamyl alcohol (24:1), and centrifuge at 10,000 rpm for 10 min. Repeat step 5 twice.

[0030] (8) Take the supernatant, add 195 μL of NaAc and 2600 μL of absolute ethanol, shake gently, and centrifuge at 10000 rpm for 5 min.

[0031] (9) Discar...

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Abstract

The invention discloses a method for reducing the content of poplar lignin with RNA interference in the field of biological technology; CCR gene segment is cloned from poplar and is used as an RNA interference segment, a constitutive promoter CaMV35S is used for promoting and expressing to construct an expression vector containing CCR gene front-back segments, agrobactrium tumefaciens is used for mediating, the front-back structure of the CCR gene is converted to the poplar, by the reactive plant PCR detection and the detection of lignin and holocellulose of transgenic poplar, the lignin content of the transgenic plant is reduced by 9.7 percent in an average way compared with the lignin content of the non-transgenic plant, the holocellulose content of the transgenic plant is improved 3.17 in an average way compared with the holocellulose content of the non-transgenic plant,, thereby providing a method for reducing the content of poplar lignin, so as to establish the basis for providing papermaking raw material in a large scale by utilizing the transgenic poplars.

Description

technical field [0001] The invention relates to a method for reducing the Klason lignin content of poplar in the field of biotechnology, in particular to a method for reducing the Klason lignin content of poplar by RNA interference. Background technique [0002] Paper is a necessity of people's life. At present, the world's pulp and paper industry mainly uses wood as the raw material for paper fiber. The basic principle of the pulp and paper process is to separate the fibers in the plant raw materials by chemical, mechanical or combined chemical and mechanical methods, and then mix them in pulp and water. The fibers are re-interwoven into a uniform and dense paper in the suspension, and the separation of the fibers is essentially a process of plasticizing or dissolving the lignin contained in the fibers. With the rapid development of the economy and the improvement of people's living standards, not only the types and quantities of paper are more and more required, but also ...

Claims

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Application Information

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IPC IPC(8): C12N15/84
Inventor 项艳魏国沈周高蔡诚宋恩慧
Owner ANHUI AGRICULTURAL UNIVERSITY
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