Method for interfering plant endogenous CLE family polypeptide hormone and antagonist polypeptide for small molecule polypeptide hormone

A technology of small molecule peptides and peptide hormones, applied in the field of genetic engineering, can solve problems such as difficult separation and detection of peptide hormones, functional research, and functional redundancy

Active Publication Date: 2013-05-08
INST OF BOTANY CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
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  • Application Information

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Problems solved by technology

However, there are still many problems in the functional research of plant polypeptide hormones using the existing methods.
For example, 1: Since the genes encoding polypeptide hormones are usually small, current gene mutation methods such as EMS mutagenesis and T-DNA insertion methods are difficult to mutate them; 2: Since polypeptide hormones function as signal molecules in plants , the content is usually very low, and it is difficult to isolate and detect these polypeptide hormones from plants with current technical means; 3: Since the genes encoding plant polypeptide hormones usually belong to a certain gene family, the encoded precursor proteins contain conserved polypeptides The coding region has redundant functions. Even if one of the members is mutated, it is difficult to see that the plant is abnormal, so it is impossible to study its function

Method used

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  • Method for interfering plant endogenous CLE family polypeptide hormone and antagonist polypeptide for small molecule polypeptide hormone
  • Method for interfering plant endogenous CLE family polypeptide hormone and antagonist polypeptide for small molecule polypeptide hormone
  • Method for interfering plant endogenous CLE family polypeptide hormone and antagonist polypeptide for small molecule polypeptide hormone

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Embodiment 1

[0031] 1. One-dimensional alanine scan of 12 amino acids of CLV3 peptide hormone

[0032] By designing primers and PCR, point mutations were introduced into the CLE domain (sequence: N-RTVPSGPDPLHH-C) of the polypeptide hormone coding region of the CLV3 gene to complete the 12 amino acid alanine (Alanine, A) of the polypeptide region. Replace, clone the PCR product (including 1857bp before the 5'end start codon ATG and 1518bp after the 3'end stop codon TGA, and 559bp genomic sequence, the nucleotide sequence of which is shown in SEQ ID No.1:) clone Into the pDONR221 vector, and then subcloned into the pBGWFS7 target vector, the resulting construction was named CLV3 RlA ,CLV3 T2A ,CLV3 V3A ,CLV3 P4A ,CLV3 S5A ,CLV3 G6A ,CLV3 P7A ,CLV3 D8A ,CLV3 P9A ,CV33 Ll0A ,CLV3 Hl1A And CLV3 Hl2A ( figure 1 ). These constructs were transformed into wild-type Arabidopsis thaliana using Agrobacterium transformation combined with inflorescence soaking, and T1 generation plants of pos...

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Abstract

The invention relates to the field of genetic engineering and in particular relates to a method for interfering plant endogenous CLE family polypeptide hormone and antagonist polypeptide for small molecule polypeptide hormone. According to the method, effective antagonist polypeptides for small molecule polypeptide hormones of CLV3, CLE1, CLE8 and CLE27 are obtained through amino acid replacement and are expressed in plants through a transgenosis method, the specific polypeptides are subjected to functional interference, the downstream signal transduction is blocked, the polypeptides are further subjected to functional research, and the potential application values on model plant Arabidopsis and crops are mined.

Description

Technical field [0001] The invention relates to the field of genetic engineering, in particular to a method for interfering with plant endogenous CLE family polypeptide hormones and an antagonistic polypeptide of small molecule polypeptide hormones. Background technique [0002] Since the discovery of systemin, the first polypeptide hormone in higher plants in the early 1990s, more than 20 small-molecule polypeptide hormones from different plants including Arabidopsis, rice, soybeans, etc. have been reported to participate in immune defense responses. , Nodule formation, cell division, stem cell maintenance and differentiation, stoma formation, anther tapetum development, pollen-stigma recognition, pollen tube guidance and petal shedding, and many other important biological processes. From the current research results, small molecule peptide hormones mainly play a role in close-range signal transduction. They interact with the receptor kinases of the LRR-RLK family on the surface...

Claims

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Application Information

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IPC IPC(8): C12N15/82A01H5/00C07K7/08
Inventor 刘春明宋秀芬国鹏任仕超徐婷婷
Owner INST OF BOTANY CHINESE ACAD OF SCI
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