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Anthurium myb transcription factor aamyb1 and its vector, engineering bacteria and application

A technology of transcription factors and genetically engineered bacteria, applied in the direction of genetic engineering, application, and use of vectors to introduce foreign genetic material, etc., can solve the problem of lack of optional genes and achieve the effect of improving the ability

Active Publication Date: 2018-04-20
ZHEJIANG XIAOSHAN COTTON & FLAX RES INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Traditional hybrid breeding must have sufficient hybrid parents and long-term experience accumulation, but with the rapid development of biotechnology, it is possible to use genetic improvement to selectively and directional breed new varieties of Anthurium
However, due to the lack of alternative genes, it has not yet been possible to obtain excellent traits through transgenic technology, and the discovery and functional verification of its important genes are the prerequisites for improving the traits of anthurium through transgenic means.

Method used

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  • Anthurium myb transcription factor aamyb1 and its vector, engineering bacteria and application
  • Anthurium myb transcription factor aamyb1 and its vector, engineering bacteria and application
  • Anthurium myb transcription factor aamyb1 and its vector, engineering bacteria and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Example 1 Obtaining the transcription factor AaMYB1 derived from Anthurium

[0020] In May 2013, the young bracts and leaves of the 6-year-old Anthurium variety "Alabama" were taken as materials, and the total RNA was extracted with the Trizol reagent produced by Invitrogen Company, and then the total RNA was purified using the reverse transcription kit produced by Promega Company. RNA is processed and reverse-transcribed to synthesize cDNA, which is used as a template. Using the homologous cloning method, design the degenerate primer pair P1-GATCTCATCATCAGGCTGCATG (SEQ ID NO: 3), P2-GATTTCCTGGGACAACAGT (SEQ ID NO: 4), amplify with the above template, the reaction program is 95 ℃ 1 min, 94 ℃ 50 s, 56 °C for 1 min, 72 °C for 1 min, 36 cycles, 72 °C for 5 min, the reagents used (LATaq enzyme) were purchased from Dalian Bao Biological Company, the amplification system: LATaq enzyme (5U / µl) 0.5 µl, 10×LAtaq buffer II 5 µl, dNTP mixture 8 µl, template 1 µl, PI 1 µl, P2 1 µl...

Embodiment 2

[0025] Embodiment 2 Preparation of recombinant genetic engineering bacteria containing transcription factor AaMYB1 coding gene

[0026]Using specific primer pair: GSPf--ctggtaccATGGAAGACGTGCCCAACCA (SEQ ID NO: 8, containing Kpn1 restriction site), GSPr-gctctagaTTAGTTTTCTGGGAATGTTAG (SEQ ID NO: 9, containing Xba1 restriction site), the high The cDNA of the first-strand product of reverse transcription was used as a template for PCR reaction. The system refers to the above-mentioned RACE reaction system. The reaction program is: 95 °C for 1 min, 94 °C for 50 s, 66 °C for 1 min, 72 °C for 1.5 min, 36 cycles, 72°C for 5 min. The obtained PCR product was recovered and subjected to Kpn1 and Xba1 double enzyme digestion, and the enzyme digestion system was carried out according to the restriction enzyme specification of TAKARA company. The gene fragment after digestion was sequenced and verified with the expression vector pCAMBIA2300 (M) recovered by the same digestion, and the sequ...

Embodiment 3

[0027] Example 3 Application of gene encoding transcription factor AaMYB1 in preparation of anthocyanin-enriched plants

[0028] The Agrobacterium strain EHA105 containing the overexpression vector prepared in Example 2 was used to infect the leaves of common tobacco aseptic seedlings, and the general leaf disk method was used to transform (materials and methods strictly refer to Horsch RB, Fry JE, Hoffman NL, Eicholtz D, Rogers SG, Fraley RT (1985) A simple and general method for transferring genes into plants. Science 227:1229–1231). After differentiation, regeneration and screening, resistant tobacco plants containing the gene encoding transcription factor AaMYB1 are obtained. After rooting, they were transplanted into the greenhouse for character identification and analysis.

[0029] After transplanting, the transgenic resistant plants all showed different degrees of color change, mainly reflected in the purple-red color of the leaves and reproductive organs, the stronger...

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Abstract

The invention discloses an anthurium myb transcription factor AaMYB1 and a carrier, engineering bacterium and application thereof. The complete prediction amino acid sequence of the transcription factor AaMYB1 is shown as SEQ ID NO:1, and encoding genes are shown as SEQ ID NO: 2. The transcription factor synthesized by MYB regulating anthocyanin is cloned from anthurium for the first time, and the ability of a target plant to synthesize the anthocyanin is improved by means of over-expression.

Description

technical field [0001] The invention relates to an anthurium myb transcription factor AaMYB1 and its carrier, engineering bacteria and application. Background technique [0002] Anthurium ( Anthurium andraeanum ) is a cosmopolitan potted and fresh-cut flower. It occupies a very important position in the cultivation of high-grade potted flowers in my country. However, like other large flowers, our country is also at a relatively backward stage in anthurium breeding. Traditional hybrid breeding must have enough hybrid parents and long-term experience accumulation, but with the rapid development of biotechnology, it is possible to use genetic improvement to selectively and directional breed new varieties of Anthurium. However, due to the lack of alternative genes, it has not yet been realized to obtain excellent traits through transgenic technology, and the discovery and functional verification of its important genes are the prerequisites for improving the traits of anthuriu...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K7/08C12N15/29C12N15/74C12N1/21C12N15/82A01H5/00A01H6/10
Inventor 马广莹朱开元邹清成刘慧春史小华张加强安霞
Owner ZHEJIANG XIAOSHAN COTTON & FLAX RES INST
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