Auxin regulation protein, coding gene thereof, and application of protein and coding gene

A technology for regulating proteins and encoding genes, which is applied in the fields of application, genetic engineering, plant genetic improvement, etc.

Active Publication Date: 2017-07-11
SHENZHEN NONGKE GROUP +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The identification of auxin transport function shows that: AtWAT1 can mediate the transport of IAA in the vacuole to the cytoplasm, but its transport process in the cell may require the regulation of corresponding regulators

Method used

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  • Auxin regulation protein, coding gene thereof, and application of protein and coding gene
  • Auxin regulation protein, coding gene thereof, and application of protein and coding gene
  • Auxin regulation protein, coding gene thereof, and application of protein and coding gene

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Example 1 Cloning of OsCOLE1 gene cDNA

[0020] 1.1 RNA extraction and DNA removal:

[0021] 1) About 200 mg of Nipponbare seedlings were taken as material, ground with liquid nitrogen, and RNA was extracted with Trizol reagent (Invitrogen).

[0022] 2) Dissolve RNA in 85 μl of water, add 10 μl of 10× buffer and 5 μl of RQ1 RNAFree DNase (1 U / μl), at 37° C. for 15 minutes to remove DNA contamination.

[0023] 3) Add 100 μl of phenol-chloroform for extraction once, take the supernatant, precipitate RNA with an equal volume of isopropanol, wash once with 70% ethanol, and dissolve in 50 μl of water.

[0024] 4) The concentration of quantitative RNA is 1 μg / μl.

[0025] 1.2 Reverse transcription:

[0026] Follow Promega's GoScript TM Reverse Transcription System (PromegaA5000) kit was carried out, and the reaction procedure was as follows:

[0027] 1) Add the following reagents in turn, mix well and react:

[0028]

[0029] 2) After the above reaction finishes, add...

Embodiment 2

[0059] The acquisition of embodiment 2 transgenic rice

[0060] 2.1 Induction culture of rice mature embryo callus

[0061] Ripe rice Nipponbare seeds are soaked in 75% ethanol for 1-2 minutes after removing the chaff. After removing ethanol, add 0.1% mercuric chloride solution to soak the seeds for 30min (shake regularly during the period). Remove the mercuric chloride solution and rinse the sterilized seeds 3-4 times with sterile water. After the seeds were placed on sterile filter paper to absorb water, they were transferred to mature embryo callus induction medium and cultured in the dark at 26°C for about 2 weeks. Carefully remove the callus grown from the scutellum of the mature embryo, and transfer it to the subculture medium of the mature embryo. Subculture under the same conditions. Calli were subcultured every two weeks. Select subcultured 5-7d calli with pale yellow color for co-cultivation.

[0062] 2.2 Preparation of Agrobacterium Transformant Infection Solu...

Embodiment 3

[0072] Identification of embodiment 3 transgenic rice

[0073] 3.1 Extraction of DNA:

[0074] 1) Cut 100mg leaves of the transgenic material, freeze them quickly in liquid nitrogen, transfer them to a mortar filled with liquid nitrogen, grind them, and transfer the powder to a 1.5ml centrifuge tube;

[0075] 2) Add 600 μl of SDS solution, mix by inverting, place in a water bath at 65°C for 60 min, and mix by inverting every 15 min);

[0076] 3) Add an equal volume of chloroform: isoamyl alcohol (V:V=24:1), mix gently by inversion, let stand for 5 minutes, and centrifuge at 12000 rpm for 15 minutes;

[0077] 4) Transfer the supernatant to another new centrifuge tube, add an equal volume of isopropanol, mix evenly by inversion, let stand for 5 minutes, and centrifuge at 12000 rpm for 15 minutes;

[0078] 5) Discard the supernatant, wash the precipitate with 75% ethanol, dry, add 30 μl sterile water to dissolve the DNA (RNase is added to the sterile water to remove RNA), and s...

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Abstract

The invention relates to an auxin regulation protein, a coding gene thereof, and an application of the protein and the coding gene. The auxin regulation protein is a protein a) composed of an amino acid residue sequence represented by sequence 2 in a sequence table, or a protein b) obtained by substituting and/or deleting and/or adding one or more amino acids to the amino acid sequence residue sequence represented by the sequence 2 in the sequence table, associating with auxin regulation and derived from the protein a). The invention also discloses the coding gene of the auxin regulation protein. The over-expression of the auxin regulation protein and the coding gene can cause the increase of the content of auxin in rice, large plants and long growth period, and the inhibitory expression of the gene causes the reduction of the content of intracellular auxin, short plants, small leaf angles and precocious flowering, so the auxin regulation protein and the coding gene participate in the regulation of plant type and flowering of plants, and have great application prospect in plant breeding.

Description

technical field [0001] The invention relates to the field of plant genetic engineering, and more specifically relates to an auxin regulatory protein OsCOLE1, a coding gene and its application. Background technique [0002] As one of the most important plant hormones, auxin (IAA) plays an important role in the growth and development of plants. As one of the most popular components in auxin-related research, the auxin transport process plays a decisive role in the process of auxin realizing its physiological effects. Compared with a large number of achievements in the study of auxin polar transport mechanism and physiological effects, there are few studies on auxin transport process in cells, so people still know little about this important plant physiological process. [0003] The most obvious effect of auxin is to promote growth, but the promotion effect on stem, bud and root growth varies with concentration. The optimum concentration of the three is stem>bud>root, a...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/415C12N15/29C12N15/84A01H5/00
CPCC07K14/415C12N15/8261C12N15/827
Inventor 王磊禹滨刘飞赵贵林张兰王维部徐妙云肖增魁
Owner SHENZHEN NONGKE GROUP
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