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Application of HsCIPK17 of Hordeum spontaneum C. Koch in Qinghai Tibet Plateau to improvement in rice resistance to abiotic stress

A wild and barley technology, applied in the field of plant genetic engineering, can solve problems such as uninformed, achieve the effect of promoting growth and development, and good application prospects

Inactive Publication Date: 2018-09-28
LANZHOU UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, there is no known CIPKs gene that has resistance / tolerance to abiotic stresses such as heavy metals and salt

Method used

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  • Application of HsCIPK17 of Hordeum spontaneum C. Koch in Qinghai Tibet Plateau to improvement in rice resistance to abiotic stress
  • Application of HsCIPK17 of Hordeum spontaneum C. Koch in Qinghai Tibet Plateau to improvement in rice resistance to abiotic stress
  • Application of HsCIPK17 of Hordeum spontaneum C. Koch in Qinghai Tibet Plateau to improvement in rice resistance to abiotic stress

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0088] Embodiment 1, seed germination and cultivation

[0089] The annual wild barley X74 (Hordeum spontaneum C.Koch) and rice Nipponbare (Oryza sativa L.ssp.japonica) seeds on the Qinghai-Tibet Plateau were surface-sterilized with 70% ethanol for 10 minutes, followed by 10% NaClO for 30 minutes, and finally rinsed with water for 8 times . The endosperm of barley and rice grains contains a large amount of nutrients, which is sufficient to meet the nutritional needs of grain germination within a week. Therefore, a simple CaCl 2 solution (0.1mM CaCl 2 ; pH 5.8) for barley and rice seed germination and seedling growth. In the dark at 25°C, sterile barley seeds were placed in CaCl 2 After one day of germination between two layers of filter paper soaked in the solution, the seeds were transferred to a new CaCl 2 The incubation was continued for 4 days on solution-soaked absorbent cotton filter paper (dark 25°C). Rice seeds were treated in the dark (4°C) for 3 days, then germi...

Embodiment 2

[0090] Embodiment 2, the cloning of HsCIPK17 gene of annual wild barley of Qinghai-Tibet Plateau

[0091] (1) Two methods used for electronic cloning of HsCIPK17. First, the known rice OsCIPK1 to OsCIPK31 ( http: / / www.ncbi.nlm.nih.gov / ; OsCIPK cDNAs gene accession numbers are as follows) full-length cDNAs as probes, search for homologous genes in barley (Hordeum vulgare L.) full-length cDNA library ( http: / / earth.lab.nig.ac.jp / ~ dclust / cgi-bin / barley_pub / ). Secondly, the conserved sequence activation loop and NAF / FISL motifs in the rice OsCIPK1 to OsCIPK31 full-length cDNAs used as probes were compared with the barley nucleotide collection (nr / nt) database ( http: / / blast.ncbi.nlm.nih.gov / ) to perform BLAST alignment of homologous fragment sequences, and perform electronic assembly and extension. Finally, the resulting full-length cDNA was analyzed with DNA STAR SeqMan and Megalign software.

[0092] OsCIPK1-OsCIPK31 cDNAs NICB Genbank accession number

[0093] ...

Embodiment 3

[0096] Example 3, Construction of the transgenic 35S::HsCIPK17 vector of the HsCIPK17 gene

[0097] 35S::HsCIPK17 ( figure 1 ). pCAMBIA2300S contains 2×CaMV 35S promoter and kanamycin resistance marker. The vector is used for the overexpression of HsCIPK17 gene in rice. Using cDNA as a template, use HsCIPK17-specific primers (Table 1) to amplify the corresponding HsCIPK17 target fragment. The PCR amplification system (50 μL system) is as follows: PCR H 2 O 17.5 μL, 2×PrimerSTAR GCBuffer 25 μL, PrimerSTAR HS DNA Polymerse 0.5 μL, dNTP mixture 4 μL, Primer F 1 μL, Primer R 1 μL, cDNA 1 μL. The PCR amplification program was as follows: pre-denaturation at 94°C for 3min, denaturation at 98°C for 10s, annealing at 65°C for 5s, extension at 72°C for 1min, 35 cycles, extension at 72°C for 10min, and storage at 16°C. Then, the target fragment was connected to the carrier through molecular cloning operations such as purification (recovery from gel cutting), enzyme digestion, and li...

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Abstract

The invention discloses application of a calcineurin B protein interaction protein kinase gene HsCIPK17 of annual Hordeum spontaneum C. Koch in the Qinghai-Tibet Plateau: the HsCIPK17 is used for constructing Oryza sativa L.; the Oryza sativa L. has heavy metal stress tolerance and also has salt tolerance and abscisic acid stress tolerance; heavy metals are mercury, cadmium and chromium; a nucleotide sequence of the gene HsCIPK17 is that the GenBank accession number is JN655677.

Description

technical field [0001] The invention belongs to the field of plant genetic engineering. Specifically, the present invention relates to the application of wild barley HsCIPK17 on the Qinghai-Tibet Plateau in improving rice resistance / tolerance to abiotic stress. Background technique [0002] In order to adapt to the changing environment, plants must respond to various biotic and abiotic stresses, thus establishing a series of regulatory mechanisms during evolution, such as sensing and decoding various stress signals, signal transduction and expression of stress-related genes. During plant growth and development, Ca 2+ Signaling is a central regulator of the physiological response of plant cells to stress (Dodd et al., 2010). Ca 2+ The change of the signal is first sensed and decoded by the calcium sensor, and then the calcium sensor interacting protein transmits the signal to the downstream, thereby activating the expression of downstream early response genes, and finally ...

Claims

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Application Information

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IPC IPC(8): C12N15/82C12N15/55A01H5/00A01H6/46
CPCC12N9/16C12N15/8271C12Y301/03016
Inventor 潘建伟潘伟槐沈金秋郑仲仲严旭
Owner LANZHOU UNIVERSITY
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