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Rheum tanguticum chalcone synthase Rt-CHS1 gene, protein and cloning method

A technology of yellow chalcone and cloning method, which is applied in the fields of genetic engineering, plant gene improvement, botany equipment and methods, etc. It can solve the problems that the chalcone synthase gene is rarely reported and has not been reported yet, and the accuracy rate is achieved. High, easy-to-operate effect

Inactive Publication Date: 2018-05-25
CHINA ACAD OF SCI NORTHWEST HIGHLAND BIOLOGY INST
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Problems solved by technology

[0004] At present, the CHS gene has been successfully cloned in petunia, orchid, Arabidopsis and other plants. There are few reports on the chalcone synthase gene of Rhubarb tangutica, and the cloning of the CHS gene and its encoded protein in Rheum tangutica has not been seen yet. Reports on Amino Acid Sequence Analysis

Method used

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  • Rheum tanguticum chalcone synthase Rt-CHS1 gene, protein and cloning method
  • Rheum tanguticum chalcone synthase Rt-CHS1 gene, protein and cloning method
  • Rheum tanguticum chalcone synthase Rt-CHS1 gene, protein and cloning method

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Embodiment 1

[0064] RNA extraction: Tangut rhubarb (Rheum tanguticum Maxim.ex Balf.) root was selected as the experimental material, and the total RNA was extracted by Cheng Xiaoli et al. (2014) improved CTAB-LiCl method. The result is as figure 1 As shown, the Nanodrop2000 micro-volume ultraviolet spectrophotometer detects its A 260 / A 280 = Between 1.9 and 2.0, indicating that the purity of the extracted RNA is high.

[0065] Synthesis of cDNA by reverse transcription: using total RNA as a template, using PrimeScript produced by TaKaRa TM 1stStrand cDNA Synthesis Kit was used for reverse transcription to obtain cDNA.

[0066] Amplification of the Rt-CHS1 gene fragment: using the cDNA as a template, according to the conserved region of the CHS gene of other Polygonaceae plants on NCBI, Primer Premier 5 software was used to design primers for PCR amplification. NO.4, shown in SEQ ID NO.5, SEQ ID NO.6 and SEQ ID NO.7, SEQ ID NO.8, the reaction system is: the total volume of PCR reactio...

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Abstract

The invention relates to a rheum tanguticum chalcone synthase Rt-CHS1 gene, a protein and a cloning method and in particular relates to the technical field of biological gene cloning. By adopting a method that an RT-PCR (Reverse Transcription-Polymerase Chain Reaction) technique and an RACE (Rapid-Amplification of cDNA Ends) technique are combined, a rheum tanguticum Rt-CHS1 gene is studied, a cDNA complete nucleotide sequence of the Rt-CHS1 gene is obtained the cloning of rheum tanguticum roots for a first time, and amino acid sequences encoded by the gene are analyzed. A base is made for further study on expression and regulation on the rheum tanguticum Rt-CHS1 gene, reference is provided for study on synthesis and regulation mechanisms of the Rt-CHS1 gene with rheum tanguticum anthraquinones, and meanwhile new background information is provided for function and evolution study on a polygonaceae gene CHS1 gene. In addition, cloning of the rheum tanguticum Rt-CHS1 gene has great significances for increasing the content of anthraquinone compounds in rheum tanguticum and improving the quality of medicinal materials.

Description

technical field [0001] The invention belongs to the technical field of biological gene cloning, and in particular relates to a Tangut rhubarb chalcone synthase Rt-CHS1 gene, protein and a cloning method. Background technique [0002] Rheum tanguticum Maxim.ex Balf. is a perennial herbaceous plant of the genus Rheum L. in the Polygonaceae family. It is distributed in eastern Tibet, Qinghai, and Gansu. It is a unique species in the Qinghai-Tibet Plateau in the bushes and half-shady slopes. Its dried roots and rhizomes are used as medicine, cold in nature and bitter in taste; it has the effects of purging and attacking stagnation, clearing heat and purging fire, cooling blood and detoxifying, expelling blood stasis and dredging menstruation, promoting dampness and reducing jaundice; it is listed in "Pharmacopoeia of the People's Republic of China" (2015) One of the three "authentic rhubarb" recorded. [0003] For Rhubarb Tangut, its chemical components include anthraquinones,...

Claims

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Application Information

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IPC IPC(8): C12N9/10C12N15/54C12N15/10
CPCC12N9/1037C12Y203/01074
Inventor 胡延萍李毅王莉王钧钱前
Owner CHINA ACAD OF SCI NORTHWEST HIGHLAND BIOLOGY INST
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