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Acid phosphatase GmPAP1-like protein gene and application thereof

A phosphatase protein and acidic technology, applied in the field of plant biology, can solve the problems of low phosphorus utilization rate, increase the risk of water pollution, phosphorus enrichment, etc., achieve good application prospects, enhance the activation and utilization of organic phosphorus, Enhanced Adaptive Effects

Active Publication Date: 2018-05-18
SOUTH CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, on acid tillage soils, the available phosphorus content is often difficult to meet the needs of crop growth (Kochian et al., 2004; von Uexküll and Mutert 1995; Vance et al., 2003)
The low utilization rate of phosphorus has become the limiting factor of modern agriculture and forestry production. In traditional agriculture, more phosphorus fertilizers are applied to meet the needs of plants for phosphorus. However, the accumulation of phosphorus in the soil not only leads to the enrichment of phosphorus in the soil, but also Increased risk of water pollution

Method used

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  • Acid phosphatase GmPAP1-like protein gene and application thereof
  • Acid phosphatase GmPAP1-like protein gene and application thereof
  • Acid phosphatase GmPAP1-like protein gene and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] The acquisition of embodiment 1 gene

[0055] 1. Hydroponic experiments to study the effect of phosphorus availability on soybean roots

[0056] Soybean variety YC03-3 was selected as plant material. After the seeds germinated, the seedlings with basically the same growth were selected and transplanted into the seedlings containing 5 μM (-P) and 250 μM (+P) KH 2 PO 4 soybean nutrient solution treatment. After 10 days of treatment, the above-ground and underground parts of soybean were harvested, and their dry weight, phosphorus content and root acid phosphatase activity were measured.

[0057] (1) Effect of phosphorus availability on soybean dry weight and phosphorus content

[0058] About 0.1 g of dry soybean sample was ground into powder and washed with H 2 SO 4 and H 2 o 2 Cook. The supernatant of each sample was then transferred to a volumetric flask and made up to 100 mL with secondary water. The phosphorus content in the solution was measured with a flow...

Embodiment 2

[0076] The construction of embodiment 2 carrier

[0077] 1. Construct an overexpression vector:

[0078] Using the soybean YC03-3 root cDNA as a template, the primer pair ORF-F and ORF-R were used to amplify the GmPAP1-like ORF1851bp fragment. After the PCR fragment was recovered and sequenced correctly, the fragment and the target vector were digested with XbaI, and the GmPAP1 The -like gene was connected to the target vector PTF101s to obtain the overexpression vector PTF101s-GmPAP1-like.

[0079] Upstream specific primer ORF-F (SEQ ID NO.7)

[0080] 5'-CCGGGGATCCTCTAGAATGATGATGAGTGGGATGG-3'

[0081] Downstream specific primer ORF-R (SEQ ID NO.8):

[0082] 5'-GCAGGTCGACTCTAGATCAAGATGCTAGTGTTGTAGCTG-3'

[0083] 2. Construction of expression vector for subcellular localization analysis:

[0084] According to conventional methods, soybean root RNA was extracted and reverse-transcribed into cDNA. Using the cDNA as a template, primers F-F and F-R were used to amplify the GmP...

Embodiment 3

[0089] The research of embodiment 3 transgenic materials

[0090] 1. Obtaining and testing of genetically modified materials

[0091] (1) Obtaining hairy root of transgenic kidney bean

[0092] The constructed overexpression vector PTF101s-GmPAP1-like and the vector pEGAD-GmPAP1-like were respectively transformed into Agrobacterium rhizogenes K599, and Agrobacterium-mediated transformation of kidney bean hairy roots was used, and this strain was used for subsequent phenotypic identification Tie.

[0093] Empty vector control: According to the above method, the overexpression vector PTF101s was empty-loaded, and the same method was used to transform the hairy root of kidney bean, and the PTF101s empty-load control line (CK) was obtained.

[0094] (2) Detection of transgenic kidney bean hairy roots

[0095] Appropriate amount of samples were taken to extract RNA after the hairy root of kidney bean was formed, and after reverse transcription into cDNA, the effect of overexpres...

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Abstract

The invention discloses an acid phosphatase GmPAP1-like protein gene from the cell wall of soybeans and application thereof. The acid phosphatase GmPAP1-like protein gene has a nucleotide sequence asshown in SEQ ID No. 1 and an amino acid sequence as shown in SEQ ID No. 2. The acid phosphatase GmPAP1-like protein gene provided by the invention is capable of participating in activation and utilization of extracellular organic phosphorus, eventually improves the low-phosphorous resistance of plants and has strong acid phosphatase activity. Overexpression of the acid phosphatase protein gene inthe roots of crops can substantially improve the organic-phosphorus activation and utilization capabilities of transgenic roots; and thus, soybean roots can be improved in adaptive capacity to low-phosphorous stress of acid soil through gene transformation, and the acid phosphatase protein gene has good application prospects in construction of transgenic soybeans with resistance to low-phosphorousstress.

Description

technical field [0001] The invention belongs to the field of plant biotechnology. More specifically, it relates to an acid phosphatase protein gene GmPAP1-like and its application. Background technique [0002] Phosphorus is an essential nutrient element for plants. It is not only the main component of cell structure, but also directly or indirectly participates in various metabolic processes of plants, such as membrane and nucleotide synthesis, photosynthesis, energy transfer and signal transduction, etc. ( Raghothama, 1999; Vance et al., 2003; Plaxton and Lambers, 2015). However, on acid tillage soils, the available phosphorus content is often difficult to meet the needs of crop growth (Kochian et al., 2004; von Uexküll and Mutert 1995; Vance et al., 2003). The low utilization rate of phosphorus has become the limiting factor of modern agriculture and forestry production. In traditional agriculture, more phosphorus fertilizers are applied to meet the needs of plants for ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/55C12N9/16C12N15/82C12N1/21A01H5/00A01H6/54
CPCC12N9/16C12N15/8271C12Y301/03002
Inventor 梁翠月吴炜炜林雁陈倩倩
Owner SOUTH CHINA AGRI UNIV
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