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Promoter regulating expression of genes in non-secreted glandular hair and application of promoter

A technology of non-secretory glandular hairs and gene regulation, applied in the field of plant biology, can solve the problems of normal plant growth burden, time and space regulation of target genes, and harm

Active Publication Date: 2016-01-27
SHANGHAI JIAO TONG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the current genetic engineering research of Artemisia annua, constitutive promoters are often used, such as the cauliflower mosaic virus 35S promoter (CaMV35S), which can drive the expression of foreign genes in all tissues and organs, and will cause excessive consumption. The substance and energy in the cell, and the expression of the target gene cannot be regulated in time and space, which is likely to cause a certain burden and harm to the normal growth of plants

Method used

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  • Promoter regulating expression of genes in non-secreted glandular hair and application of promoter
  • Promoter regulating expression of genes in non-secreted glandular hair and application of promoter
  • Promoter regulating expression of genes in non-secreted glandular hair and application of promoter

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Cloning of the Genome Sequence of the Promoter from the Artemisia annua Genome

[0033] 1. Cultivate aseptic vaccines of Artemisia annua:

[0034] Artemisia annua seeds were soaked in ethanol with a volume fraction of 75% for 1 min, then soaked in 20% (w / v) NaClO for 20 min, rinsed with sterile water for 3 to 4 times, blotted the surface moisture with sterile absorbent paper, and inoculated Cultivate on hormone-free MS solid medium at 25°C and 16h / 8h (sunlight / darkness) under light, after 14 days you can obtain sterile A. annua seedlings;

[0035] 2. Genomic DNA extraction

[0036] Put a leaf of Artemisia annua (1cm 2 About the size, take it in an ice box), add 2 steel balls. Add 300 μL TPS buffer (operate in a fume hood, TPS contains mercaptoethanol), shake at 55-60 Hz for 100 seconds. Then add 300 μL TPSbuffer (fume hood). Water bath at 65°C for 1h (shake every 20min), and the time can be extended appropriately, up to 1.5h. Cool to room temperature and centrifug...

Embodiment 2

[0052] Analyze the cis-acting elements of the AaTPS8 gene promoter to determine the type of AaTPS8 gene promoter

[0053] The length of the AaTPS8 gene promoter sequence obtained in the present invention is 1629bp. In order to find cis-acting elements on the promoter, the promoter of AaTPS8 gene was analyzed with Plantcare (http: / / bioinformatics.psb.ugent.be / webtools / plantcare / html / ). The analysis found that besides TATAbox and CAATbox, there are many cis-acting elements on the promoters cloned: ABRE, HSE, MBS, LTR, TC-richrepeats, WUN-motif, etc.

[0054] The cis-acting elements in the promoter sequence are shown in Table 4:

[0055] Table 4: Analysis of regulatory elements in the promoter sequence

[0056]

[0057] In Table 4, the positions of -1378, -691 and -1042 refer to the corresponding positions in the sequence listing listed in Example 1.

[0058] ABRE can respond to abscisic acid in plants; HSE responds to heat stress, LTR is induced by low temperature, MBS is ...

Embodiment 3

[0060] Connect the obtained promoter into the pCAMBIA-1391z vector and fuse the GUS reporter gene

[0061] In order to study the expression of the gene promoter in different plant parts, the promoter proTPS8 of the AaTPS8 gene was connected to the pCAMBIA-1391z vector and fused with the GUS reporter gene. site, the NcoI restriction site was introduced into the reverse primer, and the primer sequence is shown in the table below:

[0062] Table 5 pCAMBIA1391z-proTPS8 vector constructs PCR primers

[0063]

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Abstract

The invention discloses a promoter regulating expression of genes in non-secreted glandular hair. A nucleotide sequence of the promoter is shown as SEQ ID No:1. The invention further provides a preparation method of the promoter, a carrier with the same and an expression box. The promoter can regulate expression of target genes temporally and spatially, origination and development of the non-secreted glandular hair and metabolic regulation of secondary metabolite therein can be studied deeply, and the promoter can be applied in genetic engineering breeding for producing metabolite. Consequently, the promoter is of important significance in utilizing plant glandular hair tissue expression and genetic engineering breeding for producing the metabolite.

Description

technical field [0001] The invention relates to the field of plant biotechnology, in particular to a terpene synthase gene promoter (proTPS8) specifically expressed in plant non-secretory glandular hairs and its application in transgenic plants. Background technique [0002] Artemisia annua (Artemisia annua L.) is an annual herb of the genus Artemisia in the family Compositae. The plant has a strong volatile aroma. The essential oil of Artemisia annua extracted from the glandular hairs of Artemisia annua contains a large number of terpenoids and flavonoids with pharmacological or biocidal activity. class of compounds. There are two kinds of glandular hairs on the leaf surface of Artemisia annua: glandular secretory trichome (GST) and non-secretory glandular hairs (T-shaped trichome, TST), two kinds of glandular hairs are very important for plant growth, development, defense and pollen transmission, etc. all play a vital role. Artemisinin is a sesquiterpene lactone compound...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/113C12N15/84A01H5/00
Inventor 唐克轩陈俏沈乾付雪晴石璞孙小芬颜廷祥
Owner SHANGHAI JIAO TONG UNIV
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