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Two types of plant proteins as well as coding genes and application thereof

A technology for encoding genes and proteins, applied to two kinds of plant proteins and their encoding genes and applications, can solve the problems of cloning and functional studies of sucrose synthase encoding genes that have not been reported

Inactive Publication Date: 2013-08-14
SOUTHWEAT UNIV OF SCI & TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

So far, the cloning and functional study of the gene encoding sucrose synthase in Neosinocalamus affinis has not been reported

Method used

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  • Two types of plant proteins as well as coding genes and application thereof
  • Two types of plant proteins as well as coding genes and application thereof
  • Two types of plant proteins as well as coding genes and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] Embodiment 1, gene cloning of Cizhu MYB transcription factor and sucrose synthase

[0051] 1. Take the tender stem of Neosinocalamus affinis, extract the total RNA with Trizol reagent, and obtain cDNA by reverse transcription.

[0052] 2. Using the cDNA obtained in step 1 as a template, use specific primers MYB-F1 and MYB-R1 for PCR amplification, recover and purify the obtained PCR products, and connect them into the vector pMD18-T (purchased from Bao Biotechnology (Dalian) Co., Ltd.), obtained the intermediate vector pMD18-T-MYB, which was confirmed by sequencing. The intermediate vector pMD18-T-MYB was inserted at the TA cloning site of the vector pMD18-T with a length of 1299bp shown in Sequence 2 of the Sequence Listing A DNA fragment, the DNA fragment is named gene NaMYB1; the gene encodes a protein NaMYB1 consisting of 432 amino acid residues shown in Sequence 1 of the Sequence Listing.

[0053] 3. Using the cDNA obtained in step 1 as a template, use specific pr...

Embodiment 2

[0059] Embodiment 2, construction of recombinant vector

[0060] 1. Construction of eukaryotic bivalent recombinant plasmids

[0061] 1. Fusion of NaMYB1 and NaSuSY genes by overlapping PCR method

[0062] 1) Using the intermediate vector pMD18-T-MYB obtained in step 2 of Example 1 as a template, and MYB1 and MYB2 as primers, PCR amplification was performed to obtain DNA fragment A with the following sequence: SmaI restriction enzyme digestion sequence, sequence listing sequence 2 The full-length sequence at positions 1-1296 (excluding the stop codon tag) and the sequence at positions 1-25 of sequence 4 in the sequence listing;

[0063] 2) Using the intermediate vector pMD18-T-SuSY obtained in Step 3 of Example 1 as a template, and SuSY1 and SuSY2 as primers, PCR amplification was performed to obtain a DNA fragment B with the following sequence: SalI digestion recognition sequence, sequence listing sequence The full-length sequence of positions 1-2451 of 4, the sequence of p...

Embodiment 3

[0076] Embodiment 3, the acquisition of recombinant Agrobacterium

[0077] Take the recombinant vector pCAMBIA2301-MYB-SuSY obtained in Example 2, transform Agrobacterium tumefaciens EHA105 (purchased from Shanghai Beinuo Biotechnology Co., Ltd.) by freeze-thaw method, and obtain Agrobacterium tumefaciens EHA105 containing the recombinant vector pCAMBIA2301-MYB-SuSY , named recombinant Agrobacterium EHA105 / pCAMBIA2301-MYB-SuSY.

[0078] The recombinant vector pCAMBIA2301-MYB obtained in Example 2 was used to transform Agrobacterium tumefaciens EHA105 by the freeze-thaw method to obtain Agrobacterium tumefaciens EHA105 containing the recombinant vector pCAMBIA2301-MYB, which was named recombinant Agrobacterium EHA105 / pCAMBIA2301-MYB.

[0079] The recombinant vector pCAMBIA2301-SuSY obtained in Example 2 was used to transform Agrobacterium tumefaciens EHA105 by the freeze-thaw method to obtain Agrobacterium tumefaciens EHA105 containing the recombinant vector pCAMBIA2301-SuSY, w...

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Abstract

The invention discloses two types of plant proteins as well as coding genes and application thereof. The two types of plant proteins are derived from Neosinocalamus affinis which can be used as a good bamboo pulp papermaking material, and are respectively a protein A and a protein B, wherein the protein A is as shown by a sequence 1 in a sequence table, and the protein B is as shown by a sequence 3 in the sequence table. The protein A and a coding gene thereof can be used for regulating and controlling the cold resistance of plants, and the protein B and a coding gene thereof can be used for regulating and controlling the cellulose content of the plants. The two types of plant proteins are of great significance in cultivation of plants with strong cold resistance and high cellulose content.

Description

technical field [0001] The invention relates to two kinds of plant proteins and their coding genes and applications. Background technique [0002] With the rapid development of plant genetic engineering, the transfer of multiple genes with different functions into plant tissues and their simultaneous expression have become the focus of current research. Traditional methods of polygenic transformation such as hybridization and multiple transformations require more time and effort. The construction of multigene plant expression vectors, that is, the simultaneous insertion of multiple genes into a basic vector, is a new method that can replace traditional multigene transformation methods. [0003] In recent years, genetic engineering has expanded from genetic manipulation of structural genes to genetic manipulation of regulatory genes. Transcription factors widely exist in higher plants, which can promote the expression of various inducible genes in plants at the transcriptio...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/415C12N9/10C12N15/29C12N15/54C12N15/63C12N5/10C12N1/21C12N7/01C12N15/82C12N15/84A01H5/00
Inventor 胡尚连曹颖卢学琴段宁任鹏
Owner SOUTHWEAT UNIV OF SCI & TECH