Application of zizania sinensis ZlRc gene in increasing procyanidine content of rice seeds

A proanthocyanidin and rice seed technology, applied in the fields of application, genetic engineering, plant gene improvement, etc., can solve the problem of reduced nutritional value and achieve the effect of increasing the content of proanthocyanidin in seeds

Active Publication Date: 2022-03-25
TOBACCO RES INST CHIN AGRI SCI ACAD +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

High-precision processed rice has improved taste and digestibility, but its nutritional value has decreased

Method used

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  • Application of zizania sinensis ZlRc gene in increasing procyanidine content of rice seeds
  • Application of zizania sinensis ZlRc gene in increasing procyanidine content of rice seeds
  • Application of zizania sinensis ZlRc gene in increasing procyanidine content of rice seeds

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Embodiment 1: the acquisition of ZlRc gene

[0031] 1.1 Extraction of Chinese wild rice total RNA and preparation of cDNA

[0032] (1) Extract total RNA from Chinese wild rice:

[0033] The RNA of Chinese wild rice leaf was extracted by polysaccharide and polyphenol plant RNA extraction kit, and reverse transcribed into cDNA. The RNA extraction of Chinese wild rice leaves was carried out with reference to the instructions of the plant RNA extraction kit, and the specific experimental methods were as follows:

[0034] 1) The rice sample was rapidly ground into powder in liquid nitrogen, weighed 50 mg of liquid ammonia ground sample, added 500 μL Bufer PRL preheated at 65°C, and immediately vortexed vigorously for 60 sec.

[0035] 2) Place the lysate in a water bath at 65°C for 5 minutes, invert twice during the period, centrifuge at 12000 rpm (13400×g) for 10 minutes, transfer the supernatant to a new 1.5mL RNase-free centrifuge tube, add 0.5 times the volume of the supe...

Embodiment 2

[0052] Embodiment 2: Construction and genetic transformation of ZlRc gene overexpression vector

[0053] 2.1 Construction of ZlRc overexpression vector

[0054] The specific steps for constructing the overexpression vector are as follows: the 1390-ubi vector was digested with the restriction endonuclease BamHI, the digested product was separated by agarose gel electrophoresis, and the linearized 1390-ubi large fragment was recovered with a gel recovery kit. Using seamless clone amplification primers FP: 5'-CAGGTCGACTCTAGAGGATCCATGCACGCCATGGCCGGC-3'(SEQ ID NO:5) and FR: 5'-TTGCGGACTCTAGAGGATCCTTGATTCTTGATTCCGAAATCT-3'(SEQ ID NO:6), amplify the full-length DNA containing ZlRc The fragments were subjected to 1% agarose gel electrophoresis, and after 25 min, the target fragments were excised under ultraviolet light and purified using a gel recovery kit.

[0055] Table 2 PCR system and program

[0056]

[0057] Table 3 Seamless cloning system reaction system

[0058]

[00...

Embodiment 3

[0106] Embodiment 3: Observation of ZlRc transgenic rice and seed phenotype

[0107] The ZlRc transgenic rice and the control group Nipponbare were planted in the same greenhouse environment and given the same watering and fertilization frequency. The soil used in the test is paddy soil in southern China, and an equal amount of paddy soil is added to an equal volume of water per square box, and the test is repeated 3 times. The materials were photographed and recorded when they grew under normal water and fertilizer conditions until they started heading. When the material grows to normal firmness and yellow ripeness under normal water and fertilizer conditions, take pictures and record it. After the rice is firm and yellow, the grains and ears are separated and dried, and the chaff of the grains is manually removed.

[0108] Under the same growth conditions, ZlRc transgenic rice and control Nipponbare had roughly the same growth rate, heading and yellow ripening time. After...

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Abstract

The invention belongs to the technical field of plant genetic engineering. The invention aims to increase the procyanidine content of rice seeds, and provides application of ZlRc gene of zizania sinensis to increase the procyanidine content of the rice seeds, and the ZlRc gene has a nucleotide sequence as shown in SEQ ID No.1 or a nucleotide sequence which is formed by replacing, deleting or adding one or more nucleotides to the nucleotide sequence as shown in SEQ ID No.1 and has the same function. According to the invention, the ZlRc gene can be effectively overexpressed in rice. Under the condition that the growth environment of the ZlRc gene is consistent with that of a control plant, compared with the control group, the seeds harvested from the rice with the over-expressed ZlRc gene have higher contents of total phenols, total flavonoids and total proanthocyanidins, and higher DPPH free radical scavenging capacity and ABTS. + free radical absorption capacity.

Description

technical field [0001] The invention belongs to the technical field of plant genetic engineering, and in particular relates to the application of the Chinese wild rice wild rice ZlRc gene in increasing the content of proanthocyanidins in rice. Background technique [0002] As one of the most important food crops in my country, rice can provide essential nutrients for the human body. Rice is made into rice through cleaning, husking, rice milling and other processes. During the rice milling process, the more aleurone layer and germ are removed, the closer the remaining part is to pure endosperm, resulting in fat, protein, vitamins, minerals and other wastes. The loss of phytochemicals (such as phenolic acids, flavonoids and γ-oryzanol, etc.) is also more. Although the taste and digestibility of high-precision processed rice have been improved, its nutritional value has been reduced. With the improvement of people's living standards, consumers gradually began to pursue nutrit...

Claims

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Application Information

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IPC IPC(8): C12N15/29C12N15/84A01H5/10A01H6/46
CPCC07K14/415C12N15/825Y02A40/146
Inventor 闫宁于秀婷吴丹丹祁倩倩杨婷纪春李亚丽解颜宁张晶杜咏梅刘新民张忠锋
Owner TOBACCO RES INST CHIN AGRI SCI ACAD
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