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Gingko MADS-box transcription factor gene GbMADS9 for controlling blossoming of plants and encoding protein and application of Gingko MADS-box transcription factor gene GbMADS9

A transcription factor, plant technology, applied in the field of plant genetic engineering

Inactive Publication Date: 2017-04-26
YANGTZE UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the functions of MADS-box genes regulating flowering time and stress response in Ginkgo biloba have not been reported

Method used

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  • Gingko MADS-box transcription factor gene GbMADS9 for controlling blossoming of plants and encoding protein and application of Gingko MADS-box transcription factor gene GbMADS9
  • Gingko MADS-box transcription factor gene GbMADS9 for controlling blossoming of plants and encoding protein and application of Gingko MADS-box transcription factor gene GbMADS9
  • Gingko MADS-box transcription factor gene GbMADS9 for controlling blossoming of plants and encoding protein and application of Gingko MADS-box transcription factor gene GbMADS9

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Acquisition of Ginkgo GbMADS9 gene

[0029] S1. Extraction of total RNA from female flowers of Ginkgo biloba:

[0030] S11. Take 1 g of fresh ginkgo female flowers, fully grind them in liquid nitrogen, and transfer them to a 10 ml centrifuge tube that has been cooled, add 4 ml of CTAB extract, and take a water bath at 60°C for 30 minutes;

[0031] S12. Add 4 ml of chloroform isoamyl alcohol solution to the material obtained in step S1 and mix evenly. After ice bathing for 5 minutes, centrifuge at 4°C and 12000 rpm for 10 minutes, aspirate the supernatant, add 8M LiCl and mix evenly, at -80°C. Let stand for 60min; then centrifuge at 4°C and 12000rpm for 10min; discard the supernatant, wash twice with 70% alcohol, and invert the centrifuge tube for 10min;

[0032] S13, adding DNase to the solution treated in step S12 after digestion for 45 minutes, repeating step S12; finally adding 100 μL of DEPC water to obtain ginkgo female flower RNA.

[0033] S2. The ginkgo RNA pre...

Embodiment 2

[0040] Gene information and homology analysis of Ginkgo biloba

[0041] The length of the ginkgo GbMADS9 genome sequence of the present invention is 2820bp, the sequence contains the largest open reading of 684bp, encodes 227 amino acids, the molecular weight is 26.3kDa, and the isoelectric point is 8.5 (molecular weight, isoelectric point through the prediction website www.expasy.org / tools / protparam.htmlget). The amino acid sequence of Ginkgo biloba GbMADS9 transcription factor is 51%-65% homologous to other plant proteins (including ZMM17 from maize, AmDEFH21 from snapdragon, PhFBP24 from petunia, AtAGL32 from Arabidopsis, GGM3 from maize, TBBS of European yew and AeAP32 of Asarum) (homology analysis website http: / / www.ncbi.nlm.nih.gov), and the GbMADS9 gene has a PI motif in the C-terminal region in the amino acid residues ( FRVQPTQPNLQD), all current MADS-box transcription factors contain PI-type protein domains. (like Figure 1A shown, Figure 1A Made with DNAMAN 5.0 s...

Embodiment 3

[0045] Southern hybridization analysis of Ginkgo GbMADS9 gene: The experimental steps are carried out according to the following methods:

[0046] Extraction of DNA from ginkgo female flowers:

[0047] S1. Take 1g of fresh ginkgo female flowers, fully grind them in liquid nitrogen and transfer them to a 10ml centrifuge tube that is cold, add 4ml of CTAB extract, and take a water bath at 60°C for 30min;

[0048] S2. Add 4 ml of chloroform isoamyl alcohol solution to the material obtained in step S1 and mix evenly. After ice bathing for 5 minutes, centrifuge at 4°C and 12000rpm for 10 minutes, suck up the supernatant, add 8M LiCl and mix well, at -80°C. Let stand for 60min; then centrifuge at 4°C and 12000rpm for 10min; discard the supernatant, wash twice with 70% alcohol, and invert the centrifuge tube for 10min;

[0049] S3. Add RNase to the solution treated in step S2 and digest for 45 minutes, then repeat step S2; finally, add 100 μL of DEPC water to obtain ginkgo female fl...

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Abstract

The invention provides Gingko MADS-box transcription factor gene GbMADS9 for controlling blossoming of plants; the gene is acquired by subjecting cDNA as a template to PCR (polymerase chain reaction) amplification, wherein the cDNA is acquired by reverse transcription of total RNA of gingko female flowers. The gene has the advantages that the gene GbMADS9 is capable of controlling blossoming of plants and shortening juvenile phase of plants, and under induction of GA3, ABA, salt stress, drought stress and cold damage stress, the overexpressed gene GbMADS9 is capable of enhancing plant tolerance to osmotic stress; the gene is applicable to controlling the blossoming phase of plants.

Description

technical field [0001] The invention relates to the field of plant genetic engineering, in particular to a Ginkgo biloba MADS-box class transcription factor GbMADS9 gene that regulates plant flowering, its encoded protein and its application. Background technique [0002] Ginkgo biloba (Ginkgo biloba L.) is native to my country and belongs to the family Ginkgoaceae. Ginkgo biloba is the oldest relict plant in the world. It is known as a "living fossil" and is widely used in garden landscape, medicine, food, health care and other fields. Due to the very long childhood of Ginkgo biloba, it has brought serious obstacles to the breeding of Ginkgo biloba varieties, and many economic and social values ​​of Ginkgo biloba have been greatly restricted. In recent years, the genetic modification of flowering-related genes in woody plants by means of genetic engineering has achieved certain results in the study of shortening the childhood of woody plants. Therefore, the use of genetic...

Claims

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Application Information

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IPC IPC(8): C12N15/29C07K14/415A01H5/00
CPCC07K14/415C12N15/827C12N15/8271
Inventor 许锋杨芬叶家保张威威廖咏玲
Owner YANGTZE UNIVERSITY
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