Application of rice oslea5c‑1 gene in breeding heavy metal resistant rice varieties

A technology of anti-heavy metals and heavy metals, applied in the fields of application, genetic engineering, plant genetic improvement, etc., can solve the problems of cultivating heavy metal-resistant rice varieties that have not been seen, so as to improve the ability of plants to resist heavy metals, have good application prospects, and repair heavy metal-contaminated soil. Effect

Active Publication Date: 2018-04-20
CHONGQING UNIV
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  • Abstract
  • Description
  • Claims
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Problems solved by technology

[0004] The nucleotide sequence of the rice OsLEA5c-1 gene has been reported in the literature, but so far, there has been no report on the use of this gene in breeding rice varieties resistant to heavy metals

Method used

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  • Application of rice oslea5c‑1 gene in breeding heavy metal resistant rice varieties
  • Application of rice oslea5c‑1 gene in breeding heavy metal resistant rice varieties
  • Application of rice oslea5c‑1 gene in breeding heavy metal resistant rice varieties

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Effect test

Embodiment 1

[0030] Embodiment 1, the cloning of rice OsLEA5c-1 gene reading frame fragment

[0031] According to the OsLEA5c-1 gene candidate sequence (CT829063) found in GenBank, use the sequence processing online toolkit (SMS) biological software network http: / / www.bio-soft.net / sms to find its largest open reading frame sequence, using Primer Premier 5.0 software designed primer OsLEA5c-f: 5'-catg ccatgg taatggcgcagctggtgg-3' (SEQ ID No.1, the underlined part is the NcoI restriction site) and OsLEA5c-r: 5'-ccc ggtcacc ttagaagacgtcggagaggg-3' (SEQ ID No. 2, the underlined part is the BstEII restriction site).

[0032] Different Zhonghua No. 11 rice seedlings were taken, total RNA was extracted, and then the extracted total RNA was synthesized according to the instructions of the TaKaRa RNA PCRKit (AMV) Ver.3.0 kit, and then the first strand of cDNA was used as a template, The reading frame sequence of OsLEA5c-1 gene was amplified by primers OsLEA5c-f and OsLEA5c-r. The PCR amplifica...

Embodiment 2

[0034] Embodiment 2, the construction of rice OsLEA5c-1 gene plant expression vector

[0035] The purified PCR amplification product was double digested with NcoI and BstEII to obtain the open reading frame fragment of the OsLEA5c-1 gene, which was then connected to the backbone of the vector pCAMBIA1301 after the same double digestion to obtain the plant overexpression binary vector pCAMBIA1301-OsLEA5c- 1. The constructed overexpression binary vector pCAMBIA1301-OsLEA5c-1 was amplified with primers OsLEA5c-f and OsLEA5c-r for PCR identification, the results were as follows figure 2 shown. The results showed that the overexpression binary vector pCAMBIA1301-OsLEA5c-1 was amplified with primers OsLEA5c-f and OsLEA5c-r to obtain a 478bp DNA fragment, which was consistent with the size of the open reading frame of OsLEA5c-1 gene. At the same time, the constructed overexpression binary vector pCAMBIA1301-OsLEA5c-1 was identified by double enzyme digestion with NcoI and BstEII, ...

Embodiment 3

[0036] Embodiment 3, the preparation of rice OsLEA5c-1 gene Agrobacterium engineering strain

[0037] The overexpression binary vector pCAMBIA1301-OsLEA5c-1 was transformed into Agrobacterium tumefaciens AGLO competent cells by liquid nitrogen cold shock method, and spread on YEB solid medium (agar containing 1.5% (w / w), 50mg / L Kanamycin (Kan), 500 mg / L streptomycin (Sm) and 50 mg / L rifampicin (Rif)), pH 7.0), cultured for 2 days at 28±2°C in the dark, and challenged Take a single colony and culture it with YEB liquid medium (containing 50mg / L of Kan, 500mg / L of Sm and 50mg / L of Rif, pH7.0) for 2 days, take the bacterial solution for PCR identification, and the positive recombinant is rice OsLEA5c-1 gene Agrobacterium engineered strain, stored at -80°C for future use.

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Abstract

The invention discloses the application of the rice OsLEA5c-1 gene in cultivating heavy metal-resistant rice varieties, wherein the nucleotide sequence of the rice OsLEA5c-1 gene is shown in SEQ ID No.3, and also discloses a plant containing the rice OsLEA5c-1 gene Application of expression vectors and microbial transformants in cultivating heavy metal-resistant rice varieties; the heavy metal resistance of transgenic positive plants obtained by transforming rice OsLEA5c-1 gene into rice is significantly enhanced compared with non-transgenic rice plants, indicating that rice OsLEA5c-1 gene can be used for The cultivation of rice varieties resistant to heavy metals has good application prospects in improving rice resistance to heavy metals, promoting stable and high yields of rice.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to the application of rice OsLEA5c-1 gene in cultivating heavy metal-resistant rice varieties. Background technique [0002] Rice is one of the main food crops of mankind. More than one-half of the people in the world eat rice, and more than one-third of the population uses rice as a staple food. In today's world of food shortage, it is very important to produce enough rice to maintain enough food. If rice production is reduced, it may lead to hunger, death and social instability. With a large population, China is a big country in rice production and consumption, and rice plays a pivotal role in the national economy. However, in rice production, adversity factors such as drought, flood, low temperature, high temperature, and pests and diseases affect the quality of rice, reduce rice yield, and cause huge economic losses. Improving the stress resistance of rice, increasing ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/84A01H5/00A01H6/46
Inventor 胡廷章秦娟朱姗姗周浓何帅王贵学
Owner CHONGQING UNIV
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