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Genetic engineering application of rice gene ORYsa;SQD1

A rice and gene technology, applied in the direction of genetic engineering, application, recombinant DNA technology, etc., can solve the undisclosed Arabidopsis growth and other problems

Active Publication Date: 2019-07-19
NANJING AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0005] In previous studies, the thioisorhamnose and thioisorhamnose diacylglycerol synthase gene SQD1 in Arabidopsis is a gene induced by phosphorus deficiency, which is involved in the interaction between phospholipids and sulfolipids under phosphorus deficiency conditions. The regulatory process of homeostasis, but it has not disclosed that the gene has the function of promoting the growth of Arabidopsis

Method used

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  • Genetic engineering application of rice gene ORYsa;SQD1
  • Genetic engineering application of rice gene ORYsa;SQD1
  • Genetic engineering application of rice gene ORYsa;SQD1

Examples

Experimental program
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Effect test

Embodiment 1

[0023] Embodiment 1, the acquisition of gene sequence

[0024] AtSQD1 is a gene located on the fifth chromosome, encoding the synthesis of thioisorhamnose diglyceride synthase SQD1. The essence of SQD1 enzyme is a member of the uridine diphosphate thioisorhamnose synthase family (UDP-sulfoquinovose synthase family; SDR52E) in the short-chain dehydrogenase superfamily (Short-Chain Dehydrogenase superfamily; SDR), with NAD + The binding site can bind to ferredoxin glutamine synthase and accept sulfite (SO 3 2- ). According to the sequence of Arabidopsis SQD1 gene in NCBI, the homologous gene OsSQD1 in rice was obtained from the NCBI database, and the gene number is (same as RAB_DB) Os05g0387200. Blast and GenomeScan analysis showed that: OsSQD1 is located on the fifth chromosome of rice, the specific position is Chr5:18738602..18741791. The rice SQD1 gene contains 2 exons and 1 intron. The protein coding region of the gene is 1440bp, encoding 479 amino acids.

Embodiment 2

[0025] Embodiment 2, ORYsa; Expression pattern identification of SQD1 gene

[0026] 2.1. Extraction and transcription of total RNA to synthesize the first strand of cDNA

[0027] The rice variety "Nihonbare" was selected. After the rice seedlings grew to 10 days, they were cultured in complete nutrient solution for 1 week and then sampled (inverted one leaf, inverted two leaves, inverted four leaves, inverted second leaf sheath, inverted four leaf sheath, rhizome junction, root tip, root elongation zone and maturation zone). Rice seedlings were transplanted to potted pots when they were 30 days old, and samples were taken when the seedlings were 60 days old (flag leaves, inverted clover leaves, flag leaf sheaths, inverted clover leaf sheaths, stalks, stem knots and panicles). The total RNA was extracted with TriZol reagent, and the quality of the total RNA was identified by agarose gel electrophoresis, and then the RNA content was determined on a spectrophotometer. The obtai...

Embodiment 3

[0036] Example 3, ORYsa; SQD1 overexpression material acquisition and functional identification

[0037] 3.1 Construction of OsSQD1 overexpression vector

[0038] In order to construct the overexpression material of OsSQD1, we chose the vector pTCK303 commonly used in our laboratory as the expression vector ( Figure 9 ). According to the cDNA sequence of OsSQD1 published on the NCBI website, use DNAMAN to perform restriction enzyme analysis, and select KpnI and SpeI that are not in the cDNA and have multiple cloning restriction sites in the vector, and the target fragment can be directly combined with the pTCK303 vector. Enzyme cleavage and enzyme ligation. The primers are shown in the table (Table 1). The full-length cDNA sequence fragment of OsSQD1 was obtained by PCR amplification using high-fidelity PCR Mix using the cDNA of Nipponbare leaves as a template. Purify and recover the PCR product, and connect it into the cloning vector pEASY-Blunt. After sequencing to con...

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Abstract

The invention discloses genetic engineering application of rice phosphorus-deficiency response gene ORYsa;SQD1, including application of the gene in improving the absorption and utilization efficiencyof phosphorus in soil, increasing rice yield and influencing the seed developmental capacity, as well as a cDNA sequence SEQ ID NO.1 of the rice phosphorus-deficiency response gene ORYsa;SQD1 and anamino acid sequence SEQ ID NO.2 coded by the gene. The gene is the first report in monocotyledons, and achieves an important effect in the aspect of improving the absorption and utilization efficiencyand the yield of phosphorus in soil.

Description

technical field [0001] The invention relates to the technical field of plant genetic engineering, and relates to the genetic engineering application of rice gene ORYsa; SQD1. Background technique [0002] Rice is widely planted all over the world. It is the most important food crop in Asia, and it is also one of the main food crops in my country. The rice planting area in China accounts for 1 / 3 of the global cereal crop planting area, and rice production accounts for 10% of the country's total grain production. 50%, is the most important crop to ensure my country's food security (Hu Peisong et al., 2002). [0003] Phosphorus is one of the macronutrient elements needed for plant growth, and it is also one of the easily deficient nutrients in many soils. Although total phosphorus content in soils is often high, phosphorus availability is the key factor limiting plant growth and productivity. This contradiction occurs because the average concentration of soluble phosphorus in ...

Claims

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Application Information

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IPC IPC(8): C12N15/82A01H5/00A01H6/46
CPCC12N9/0006C12N15/8261C12N15/8242
Inventor 孙淑斌孙雅菲王小文
Owner NANJING AGRICULTURAL UNIVERSITY
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