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Method for cultivating transgenic plant with reduced anthocyanin content and delayed flowering time

A technology for transgenic plants and anthocyanin content, which is applied in the field of cultivating transgenic plants with reduced anthocyanin content and delayed flowering time, and can solve problems such as unclear molecular mechanism

Active Publication Date: 2021-11-12
THE INST OF BIOTECHNOLOGY OF THE CHINESE ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the molecular mechanism of regulation of flowering time of leguminous forages is still unclear and needs further research

Method used

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  • Method for cultivating transgenic plant with reduced anthocyanin content and delayed flowering time
  • Method for cultivating transgenic plant with reduced anthocyanin content and delayed flowering time
  • Method for cultivating transgenic plant with reduced anthocyanin content and delayed flowering time

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0097] Embodiment 1, the cloning of EAF2 gene

[0098] 1. Phenotype analysis of mutant eaf2

[0099] Take 30 mutant eaf2 seeds or Medicago truncatula R108 seeds, vernalize them at 4°C for 7 days, plant them in nutrient soil, and culture them in alternating light and dark (16h light / 8h dark), observe the phenotype and count.

[0100] See the growth status of some mutants eaf2 and Medicago truncatula R108 in alternating light and dark culture for 40 days. figure 1 Middle A (WT is Medicago truncatula R108, eaf2 is mutant eaf2). The results showed that after 40 days of alternating light and dark culture, the mutant eaf2 had flowered, but Medicago truncatula R108 had not flowered.

[0101]Count the flowering time and the number of nodes of each strain of Medicago truncatula (that is, the number of days of growth and the number of nodes of Medicago truncatula when the first flower fully opened), and then take the average value. see results figure 1 Middle B (WT is Medicago tru...

Embodiment 2

[0121] Embodiment 2, acquisition of complementary strains, phenotypic identification and detection of anthocyanin content

[0122] 1. Construction of recombinant plasmid 35S::EAF2

[0123] 1. Using the pMDC83 vector as a template, PCR amplification was performed with a primer pair composed of primers caccGFP-F: 5′-caccATGGGTAAAGGAGAACTTTTCA-3′ and primer HA-R+6*Gly: 5′-GCCACCCCCTCCGCCACCGGCATAATCAGGCACATCG-3′ to obtain about 800 bp GFP fragment.

[0124] 2. Using the PCR amplification product obtained in Step 3 of Example 1 as a template, use a primer pair consisting of primer 6*Gly-NFY-B4-F: 5′-GGTGGCGGAGGGGGTGGCATGGCTGAAACGGAGGAG-3′ and primer MtNF-YB4-R. PCR amplified to obtain about 420bp NF-YB4 fragment.

[0125] 3. The GFP fragment and NF-YB4 fragment were mixed as a template, and a primer pair composed of primer caccGFP-F and primer MtNF-YB4-R was used for PCR amplification to obtain a GFP-NF-YB4 fragment of about 1200 bp.

[0126] 4. The GFP-NF-YB4 fragment and the ...

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Abstract

The invention discloses a method for cultivating a transgenic plant with reduced anthocyanin content and delayed flowering time. An amino acid sequence of an EAF2 protein is as shown in SEQ ID NO: 2. Experiments prove that a recombinant plasmid 35S:: EAF2 is introduced into a mutant eaf2 to obtain a complementary plant; and compared with the mutant eaf2, the content of anthocyanin in leaves of the complementary plant is remarkably reduced, the number of flowering nodes is increased, and the flowering time is delayed. Therefore, the EAF2 protein can be used for regulating and controlling anthocyanin synthesis, flowering node number and flowering time. The method has an important application value.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for cultivating transgenic plants with reduced anthocyanin content and delayed flowering time. Background technique [0002] Plant flowering marks the transition from the vegetative stage to the reproductive stage, and plays an important role in the reproduction of plant offspring and ecological adaptation; the date of flowering of crops has an important impact on the high yield, stable yield, quality and regional distribution of crops. Breeders regard suitable flowering time as one of the key agronomic traits, and have been working hard to obtain excellent germplasm with ideal flowering time. The analysis of plant flowering time has always been a research hotspot in many disciplines such as plant molecular genetics, evolutionary biology, ecology and crop breeding. At present, the molecular mechanism of regulation of flowering time of leguminous forages is still...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/415C12N15/29C12N15/82A01H5/02A01H6/54
CPCC07K14/415C12N15/825C12N15/827
Inventor 林浩季文楷牛丽芳
Owner THE INST OF BIOTECHNOLOGY OF THE CHINESE ACAD OF AGRI SCI
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