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Russian dandelion drought-tolerant gene TkMYC2, protein, primer, vector, host bactera and application of Russian dandelion drought-tolerant gene TkMYC2

A rubber grass and protein technology, applied in application, genetic engineering, plant genetic improvement and other directions, can solve the problems of rubber yield reduction, economic loss, and drought-resistant functions that have not yet been reported, and achieve the goal of improving drought resistance and improving drought resistance. Effect

Active Publication Date: 2020-09-15
SHIHEZI UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, annual droughts cause severe reductions in rubber production, causing huge economic losses
[0003] TkMYC2 gene is a drought tolerance gene of rubber grass, the drought tolerance of rubber grass transfected with TkMYC2 gene is greatly improved, but the drought tolerance function of this gene has not been reported yet

Method used

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  • Russian dandelion drought-tolerant gene TkMYC2, protein, primer, vector, host bactera and application of Russian dandelion drought-tolerant gene TkMYC2
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  • Russian dandelion drought-tolerant gene TkMYC2, protein, primer, vector, host bactera and application of Russian dandelion drought-tolerant gene TkMYC2

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0026] Obtaining Transgenic TkMYC2 Gene Rubber Grass

[0027] (1) Extraction of rubber grass total RNA

[0028] Total RNA was extracted using an RNA extraction kit (EasyPure Plant RNA Kit), which was purchased from Beijing Quanshijin Biological Co., Ltd.

[0029] 1) Grind 0.5 g of different tissue samples quickly and fully into powder in liquid nitrogen, add 1 ml of BB6 and 10 μL of β-mercaptoethanol (pre-mixed), and vortex vigorously to mix. Stand at room temperature for 3 minutes.

[0030] 2) Centrifuge at 12 000rpm for 4min, carefully pipette the supernatant from the centrifuge tube into an RNase-free centrifuge tube

[0031] 3) Add 0.5 times the volume of absolute ethanol (pre-cooled at 4°C) to the supernatant, and mix well.

[0032] 4) Vortex to mix and disperse the precipitate.

[0033] 5) Add the precipitation mixture into a centrifuge tube, centrifuge at 12 000 rpm for 30 s, and discard the effluent.

[0034] 6) Add 500 μL of CB6, centrifuge at 12 000 rpm for 30 s...

Embodiment 2

[0080] PEG-induced expression analysis

[0081] The wild-type rubber grass aseptic seedlings grown for three months were subjected to hydroponic seedling hardening for three days, and the seedling hardening method was as in reference (De novo Transcriptome Sequencing of MeJA-Induced Taraxacum koksaghyz Rodinto Identify Genes Related to Rubber Formation). The wild-type rubber grass was treated with 20% PEG to completely submerge the roots. The PEG induction time gradient was set to 0, 6, 12, and 24 hours, and the rubber grass root tissues (1cm below the leaf base) were collected respectively, and the same parts were kept as much as possible; three biological replicates were immediately frozen in liquid nitrogen and placed in a -80°C refrigerator. save. Extraction of root total RNA from different samples using RNA Extraction Kit (EasyPure Plant RNA Kit) and reverse transcription kit (EasyScript One-Step gDNA Removal Kit and cDNA Synthesis Supermix) with Anchored Oligo(dT)18 pri...

Embodiment 3

[0085] Identification of Transgenic TkMYC2 Gene Rubber Grass Plants

[0086] The transgenic TkMYC2 gene rubber grass in Example 1 was hardened indoors for 2 days, the excess medium at the root was washed, and transplanted into nutrient soil. The nutrient soil is composed of vermiculite and humus in a ratio of 1:2. In the cultivation room, cultivate under normal conditions for about 20 days, and extract the DNA of the candidate transgenic rubber grass according to the instructions of the kit (EasyPure Plant Genomic DNA Kit).

[0087] 1) Take 0.5 g of rubber grass leaf tissue, add liquid nitrogen and grind it thoroughly.

[0088] 2) Add 250 μL RB1 and 15 μL Rnase A to each centrifuge tube.

[0089] 3) Add the sample into the centrifuge tube in step 2, mix well, and put it in a water bath at 55°C for 15 minutes.

[0090] 4) Centrifuge at 12000 rpm for 5 minutes, and absorb the supernatant.

[0091] 5) Add 100 μL of PB1, mix well, bathe in ice for 5 minutes, and centrifuge at ...

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Abstract

The invention relates to the technical field of biology, in particular to a Russian dandelion drought-tolerant gene TkMYC2, protein, primers, a vector, host bacteria and application of the Russian dandelion drought-tolerant gene TkMYC2. The nucleotide sequence of the cDNA of the TkMYC2 gene is as shown in SEQ ID NO: 1. The amino acid sequence of the protein encoded by the gene is shown as SEQ ID NO: 2. Under the stress of natural drought, compared with a wild type, the leaves of transgenic Russian dandelion with overexpression of TkMYC2 is good in growth condition. Overexpression of the TkMYC2gene can obviously improve the enzyme activity of POD, SOD and CAT to respond to drought pressure. The Russian dandelion TkMYC2 gene provides a new solution for improving the drought resistance of Russian dandelion.

Description

technical field [0001] The invention relates to the field of biological technology, in particular to a rubber grass drought-tolerant gene TkMYC2, a protein, a primer, a carrier, a host bacterium and applications thereof. Background technique [0002] Rubber grass (Taraxacum kok-saghyz Rodin), also known as Russian dandelion, is a perennial diploid rubber-producing plant. Studies have found that the physical and chemical properties of the rubber produced by rubber grass are similar to those produced by rubber trees, and the root tissue of rubber grass has up to 20% dry rubber. However, the annual drought will lead to a severe reduction in rubber production, resulting in huge economic losses. [0003] TkMYC2 gene is a drought tolerance gene of rubber grass, and the drought tolerance of rubber grass transfected with TkMYC2 gene is greatly improved, but the drought tolerance function of this gene has not been reported yet. Contents of the invention [0004] One of the object...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/415C12N15/84C12N15/29C12N15/11
CPCC07K14/415C12N15/8273C12N15/8205
Inventor 闫洁黄俊张云川孙军亭孙辉
Owner SHIHEZI UNIVERSITY
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