Gene for adjusting plant luminous energy application and oil and fat accumulation and application thereof

A gene and photosynthesis technology, applied in the field of genetic engineering, can solve the problems such as gene reports of photosynthesis and oil accumulation in oil crops seeds that have not been found, so as to improve photosynthesis and oil accumulation capacity, increase plant yield and oil content, The effect of improving related traits

Inactive Publication Date: 2010-06-23
ZHEJIANG ACADEMY OF AGRICULTURE SCIENCES
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] Although the research on plant photosynthesis and oil synthesis has attracted much attention, functional genes related to these two important biological processes have been cloned one after another, but so far, no one has found the ability to simultaneously regulate photosynthesis and oil accumulation in oil crop seeds gene reporter

Method used

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  • Gene for adjusting plant luminous energy application and oil and fat accumulation and application thereof
  • Gene for adjusting plant luminous energy application and oil and fat accumulation and application thereof
  • Gene for adjusting plant luminous energy application and oil and fat accumulation and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Example 1. Cloning of transcription factor gene BnWRI1 regulating plant light energy utilization and oil accumulation

[0045] 1.1 Extract the total RNA of Brassica napus immature seeds with Trizol reagent (purchased from Invitrogen Company), and use oligo-d(T) 18 As a primer, use M-MLV reverse transcriptase (Promega Company) to reverse-transcribe the obtained RNA into cDNA, use the synthesized cDNA as a template, and use the primer pair BnWRI1-1 (5'-ATGAAGAGACCCTTAACCACTTC-3') and BnWRI1 -2 (5'-CCTTCAGACAGAATAGTTCCAAG-3') was amplified by PCR.

[0046] The cDNA was amplified with LA-Taq (Dalian TaKaRa Company), and the PCR reaction conditions were:

[0047] 94°C, 5min, 1 cycle; 94°C, 30sec, 58°C, 40sec, 72°C, 90sec, 33 cycles; 72°C, 10min.

[0048] The PCR amplified product was connected to the carrier pMD18-T (Dalian TaKaRa Company) to obtain a recombinant plasmid containing the amplified fragment, which was named pMD-BnWRI1. After sequencing, the obtained PCR produ...

Embodiment 2

[0051] Example 2, Construction of BnWRI1 seed-specific expression vector NAPIN Pr::BnWRI1

[0052] 2.1 Firstly, using rapeseed genomic DNA as a template, use primers to perform PCR amplification on napin5 (5'-AAGCTTTCTTCATCGGTGATTGA-3') and napin3 (5'-TCGTGTATGTTTTTAATCTTGTTTG-3') to obtain the rapeseed napin promoter ( AF420598 ), the DNA fragment containing the 35S promoter between the restriction sites EcoRI-BamHI in the plant expression vector pFGC5941 was replaced with the promoter fragment to obtain the intermediate vector pFGC-NAPIN. The DNA fragment encoding BnWRI1 was excised from the plasmid pMD-BnWRI1 by double digestion with BamHI and EcoRV, and connected with the recovered pFGC-NAPIN vector fragment after double digestion with BamHI and SmaI.

[0053] 2.2 CaCl for ligation products 2 Transform Escherichia coli (E.coli) DH5α competent cells with 50mg / L kanamycin-containing LB resistance plates, and positive clones were amplified by PCR with primer pairs BnWRI1-1 ...

Embodiment 3

[0054] Embodiment 3, the acquisition of BnWRI1 transgenic Arabidopsis

[0055] 3.1 The seed-specific expression vector NAPIN Pr::BnWRI1 constructed in Example 2 was transformed into Agrobacterium tumefaciens EHA105 by the heat shock method and placed on the LB resistance plate containing 50 mg / L kanamycin and 25 mg / L rifampicin Cultured at 28°C. Single clones were picked and verified by PCR for plant transformation.

[0056] A single colony of Agrobacterium containing the plasmid NAPIN Pr::BnWRI1 was picked and inoculated into 5 ml of YEP culture solution containing 50 mg / L kanamycin and 25 mg / L rifampicin, and cultured at 28°C with shaking at 150 rpm for 2 days. According to the ratio of 1:50, expand the culture in fresh YEP liquid medium to the logarithmic growth phase (bacterial solution OD 600 ≈1.5). Centrifuge at 3000rmp for 10min, discard the supernatant. The pellet was resuspended in osmotic medium, (composed of 1 / 2MS liquid medium containing 5% sucrose, 44mM 6-benz...

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Abstract

Belonging to the field of gene engineering, the invention relates to an AP2 / EREBP transcription factor bNwrI1 gene and an application thereof in adjusting plant seed photosynthesis and oil and fat anabolism. The transcription factor is extracted from brassica napus and is one of the following polypeptides: a polypeptide provided with an amino acid sequence as SEQ ID NO:2 indicates, a polypeptide which is formed with the amino acid sequence replaced, deleted and added by one or more than one amino acid residue and provided with the amino acid sequence and a polypeptide with functions thereof derivating from the polypeptide provided with the amino acid sequence. The transcription factor and coding genes thereof enjoy extensive application value in plant culturing field and can enable the plant to coordinately regulate photosynthesis and fatty acid anabolism, improves seed photosynthesis and oil and fat accumulation capacity, thus improving plant yield and oil content as well as correlated characteristics.

Description

technical field [0001] The invention belongs to the field of genetic engineering and relates to an AP2 / EREBP transcription factor BnWRI1 gene and its application in regulating plant seed photosynthesis and oil synthesis and metabolism. Background technique [0002] In addition to its important nutritional value to humans, vegetable oils are also used as raw materials for a wide range of non-edible products, especially as global energy is becoming increasingly tense, and the use of vegetable oils to produce renewable energy has become an important strategic choice for countries around the world to deal with energy crises. Increasing oil production per unit area is the goal that oil crop breeding has always pursued. Oil content and yield per unit area are two components of oil production per unit area, and are also important determinants of oil crop production efficiency (Li Yunchang et al., Chinese Journal of Oil Crops 28:92-96, 2006). [0003] The economic trait of seed oil...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K14/415C12N15/29C12N15/63C12N5/10C12N15/11C12N15/82A01H1/00A01H1/02A01H5/00
Inventor 薛红卫黄锐之吴学龙邢梅青胡张华许永汉何海燕柯丽萍
Owner ZHEJIANG ACADEMY OF AGRICULTURE SCIENCES
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