Key gene PeSCE1 for controlling formation of adventitious roots of poplars and plant height development and application of the key gene PeSCE1

A key gene, poplar technology, applied to the key gene PeSCE1 that controls the formation of adventitious roots and plant height development of poplar and its application field, to achieve the effect of reducing plant height

Inactive Publication Date: 2020-08-21
YANGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the key enzyme gene SCE1 in the SUMOylation process is still less

Method used

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  • Key gene PeSCE1 for controlling formation of adventitious roots of poplars and plant height development and application of the key gene PeSCE1
  • Key gene PeSCE1 for controlling formation of adventitious roots of poplars and plant height development and application of the key gene PeSCE1
  • Key gene PeSCE1 for controlling formation of adventitious roots of poplars and plant height development and application of the key gene PeSCE1

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0024] Example 1 Cloning of PeSCE1 gene by RACE technology

[0025] Based on the results of the applicant's previous research on the expression profile chip of poplar adventitious roots, Oligo 6 was used to design the RACE primers at the 3' end and perform 3' RACE to obtain the 3' cDNA end fragment, which was cloned into a T-vector, and the insert fragment was screened by PCR. Sequencing, Blast confirmed that the above fragments were homologous to genes related to other plants. In the same way, design the RACE primers at the 5' end according to the obtained correct 3' cDNA end fragment sequence, perform 5' RACE, obtain the 5' cDNA end fragment, clone it into T-vector, and perform PCR screening on the insert fragment. sequencing.

[0026] Primer sequence:

[0027]

[0028] 3'RACE reaction procedure

[0029] (1) For reverse transcription, add the following components to an RNase-free centrifuge tube placed on ice:

[0030]

[0031] (2) Mix gently, centrifuge briefly, a...

Embodiment 2

[0085] Embodiment 2PeSCE1 gene plant expression vector construction

[0086] The overexpression vector of PeSCE1 gene was constructed by Gateway cloning technology. Using specific PCR primers (PeSCE1 ORF primers in Example 1), cDNA was used as a template for PCR amplification, and the PeSCE1 gene ORF was constructed into an entry vector. The entry vector is pCR TM 8 / GW / TOPO TM vector (Invitrogen). The reaction system is: Fresh PCR product (purified) 10-20ng; Salt solution 1μl; pCR TM 8 / GW / TOPO TM vector 1μl; add sterile ddH 2 O to make up 6 μl. The reaction procedure is: stand at room temperature for 30 min.

[0087] The positive clones were picked from the screening culture plate for PCR detection and sequencing verification, and the entry vector with PeSCE1 gene was subjected to LR reaction with the plant expression vector pBI121. Vector plasmid such as figure 1 shown. The reaction system is: linearized dentry clone 100ng; purified destination vector (100ng / μl) 1...

Embodiment 3

[0088] The genetic transformation of embodiment 3PeSCE1 gene

[0089] The constructed 35S::PeSCE1 overexpression vector was transformed into Agrobacterium strain EHA105 by liquid nitrogen freeze-thaw method, and the PeSCE1 gene was transferred into poplar by Agrobacterium. Experimental results such as Figure 1~4 As shown, among them, figure 1 It is a structural schematic diagram of the constructed plant expression vector 35S::PeSCE1; figure 2 Molecular detection of transgenic poplar overexpressing PeSCE1 gene; image 3 It is the overall morphological comparison between the transgenic poplar overexpressing the PeSCE1 gene and the non-transgenic poplar; Figure 4 It is the number of adventitious roots of the transgenic poplar overexpressing PeSCE1 gene and the non-transgenic poplar. From the results, it can be clearly concluded that the number of adventitious roots and plant height of the transgenic poplar overexpressing the PeSCE1 gene were significantly reduced, indicati...

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Abstract

The invention discloses a key gene PeSCE1 for controlling formation of adventitious roots of poplars and plant height development and application of the key gene PeSCE1, wherein the nucleotide sequence of the key gene PeSCE1 is shown as SEQ ID NO.1, and the amino acid sequence of an expression protein of the key gene PeSCE1 is shown as SEQ ID NO.2. The invention further discloses application of the key gene PeSCE1. The method comprises the following steps: cloning a PeSCE1 gene through an RACE technology by taking adventitious roots of poplar Nanlin 895 as a material, and constructing an overexpression vector 35S:: PeSCE1 by utilizing a Gateway technology, and transferring the overexpression vector 35S:: PeSCE1 into the poplar. The number of adventitious roots of the PeSCE1 transgenic poplar is obviously reduced, and the plant height is also obviously reduced. Therefore, the PeSCE1 gene is a key gene for regulating and controlling formation of adventitious roots and plant height development of poplars, and has important application value in the fields of forest genetic engineering and clone forestry.

Description

technical field [0001] The invention belongs to the technical field of plant genetic engineering, in particular to a key gene PeSCE1 for controlling the formation of adventitious roots and the development of plant height of poplar and its application. Background technique [0002] Poplar is one of the important fast-growing industrial timber species and one of afforestation species, which plays an important role in the sustainable development of clonal forestry. In the process of raising seedlings, poplars are usually raised by cuttings, so the difficulty of rooting cuttings directly affects the survival rate of afforestation, and is related to the adaptability and stress resistance of the clones. Although most poplars are relatively easy to take root from cuttings, there are still many good clones that are difficult to take root from cuttings. The importance of tree height to timber tree species is self-evident, and it is an important trait for breeders to improve varietie...

Claims

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Application Information

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IPC IPC(8): C12N15/52C12N9/00C12N15/82A01H5/06A01H6/00
CPCC12N9/00C12N15/8261
Inventor 刘思安胥猛祁浩然吴玲
Owner YANGZHOU UNIV
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