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A kind of Lycoris llbhlh19 gene and its expressed protein and application

A long tube lycoris and gene technology is applied in the field of genetic engineering to achieve the effects of improving ornamental characters and promoting accumulation

Active Publication Date: 2021-12-07
DAQIAN ECOLOGY & LANDSCAPE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Gene regulation of anthocyanin biosynthesis pathways is very widespread in plants, but there are few reports on the transcription factors related to the regulation of flower color in Lycoris longifolia, and the corresponding expression regulation mechanism is also poorly understood. functions in regulation, which may provide useful molecular tools for improving ornamental traits in plants using genetic engineering

Method used

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  • A kind of Lycoris llbhlh19 gene and its expressed protein and application
  • A kind of Lycoris llbhlh19 gene and its expressed protein and application
  • A kind of Lycoris llbhlh19 gene and its expressed protein and application

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Experimental program
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Effect test

Embodiment 1

[0028] Example 1: Cloning and expression analysis of LlbHLH19 gene

[0029] (1) Clone LlbHLH19 gene

[0030] In this example, TIANGEN Plant RNA Extraction Kit (DP432) was used to extract total plant RNA. Using TaKaRa PrimeScript TM The RT Master Mix (Perfect Real Time) reverse transcription kit reverse-transcribed the extracted RNA into cDNA, and the final cDNA was diluted 20 times with water as a template for real-time quantitative PCR and subsequent gene cloning.

[0031] The LlbHLH19 gene that may be involved in the synthesis of anthocyanins in Lycoris longifolia was screened according to the Lycoris longifolia transcriptome database, and specific primers were designed using the software Oligo 7. The primer sequence is:

[0032] F: 5'-CACATTCTCAAAATAAACCCTC-3';

[0033] R: 5'-GTCGTAGCAAAACAAGAAGCAC-3'.

[0034] Lycoris cDNA was used as a template, and F and R were used as primers, and PCR amplification was carried out in a 20 μL reaction system. The amplification prog...

Embodiment 2

[0042] Example 2: Vector construction and functional verification of LlbHLH19

[0043] I. Observation of subcellular localization of LlbHLH19 gene expression

[0044] (1) Cloning and sequencing of the target gene LlbHLH19

[0045] The LlbHLH19 gene ORF sequence was constructed between the two restriction sites Sal I and KpnI of the pCAMBIA1300-GFP vector, and the forward and reverse primers containing the restriction sites were designed using CE design as follows:

[0046] F: 5'-AAGCTTCTGCAGGGGCCCGGGATGGATCCGAATTCTCTCGTCTC-3';

[0047] R: 5'-CACTAGTATTTAAATGTCGACTGTGACTCTCCCAAGAGCTCCA-3'.

[0048]Lycoris cDNA was used as a template, and F and R were used as primers, and PCR amplification was carried out in a 20 μL reaction system. The amplification program was: 94°C for 4min; 35 cycles of 94°C for 30s, 58°C for 30s, and 72°C for 2min; 72°C for 10min. The obtained PCR products were subjected to agarose electrophoresis, gel-excision recovery and clone sequencing.

[0049] (...

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Abstract

The invention discloses a Lycoris lycoris LlbHLH19 gene and its expressed protein and application, belonging to the field of plant molecular biology. The nucleotide sequence of the LlbHLH19 gene is shown in SEQ ID NO.1, and the amino acid sequence of the LlbHLH19 gene is shown in SEQ ID Shown in NO.2. bHLH is an important transcription factor in plant anthocyanin biosynthesis, its expression level is highest at the early bud stage of Lycoris, and its expression level gradually decreases at the middle and large bud stages. Overexpressing the LlbHLH19 gene of Lycoris in Nicotiana early annulus can change the flower color of Nicotiana early annulus from pink to dark red, and it is a transcription factor that can change the flower color. Applying the LlbHLH19 gene to the transgenic plant can improve the flower color of the plant so as to enhance its ornamental value, and has practical application value.

Description

technical field [0001] The invention belongs to the technical field of genetic engineering, and more specifically relates to a Lycoris lycoris LlbHLH19 gene and its expressed protein and application. Background technique [0002] Lycoris longituba (Lycoris longituba) belongs to the family Amaryllidaceae (Lycoris) and is endemic to my country, mainly distributed in Jiangsu and Anhui provinces. Because of its beautiful flower shape and rich flower colors, it is widely used in fresh cut flowers, potted plants and garden landscape applications, and has high development potential and utilization value. And bHLH is an important transcription factor in the process of plant anthocyanin biosynthesis. At present, bHLH genes that regulate the transcriptional regulation of anthocyanin biosynthesis in many flower plants have been cloned and functionally identified. Gene regulation of anthocyanin biosynthesis pathways is very widespread in plants, but there are few reports on the transcr...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/29C07K14/415C12N15/84C12N1/21A01H5/02A01H6/82
CPCC07K14/415C12N15/8205C12N15/825
Inventor 杨秀莲宰舟颖王欣刘华岐岳远征王良桂施婷婷李小婷刘芳伊王明珠
Owner DAQIAN ECOLOGY & LANDSCAPE
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