Scientia Agricultura Sinica ›› 2014, Vol. 47 ›› Issue (12): 2292-2299.doi: 10.3864/j.issn.0578-1752.2014.12.002

• CROP GENETICS & BREEDING·GERMPLASM RESOURCES·MOLECULAR GENETICS • Previous Articles     Next Articles

Isolation and Functional Analysis of Salt-Responsive Gene TaSRP in Wheat

 HU  Di-1, 2 , XU  Zhao-Shi-2, CUI  Xiao-Yu-2, CHEN  Ming-2, LI  Lian-Cheng-2, MA  You-Zhi-2, ZHANG  Xiao-Hong-1   

  1. 1、College of Agronomy, Northwest A&F University/State Key Laboratory of Crop Stress Biology for Arid Areas, Yangling 712100, Shaanxi;
    2、Institute of Crop Sciences, Chinese Academy of Agricultural Sciences (CAAS)/National Key Facility for Crop Gene Resources and Genetic Improvement/Key Laboratory of Biology and Genetic Improvement of Triticeae Crops,Ministry of Agriculture, Beijing 100081
  • Received:2013-12-16 Online:2014-06-15 Published:2014-03-03

Abstract: 【Objective】 Salt is one of the major constraints to yield and quality of wheat in China. How to improve salt tolerance of crops has become an important objective of breeding. The study provided experimental data for further study of the function and the mechanism of TaSRP gene. 【Method】 TaSRP was obtained by analysis of salt treatment wheat de novo transcriptome assembly. Homologous sequences of TaSRP in O. sativa, Arabidopsis and Glycine max were selected using the NCBI BLAST analysis program. Homology analysis and multiple alignments were performed with DNAMAN and MEGA5.05. Real-time PCR of TaSRP specific primers was used to analyze the expression patterns under various abiotic stresses. The coding region of TaSRP was fused to the N-terminal end of GFP under control of the CaMV 35S promoter as 16318hGFP-TaSRP. The 16318hGFP- TaSRP and control vectors were transformed into Arabidopsis protoplasts for subcellular localization. Results were visualized by a confocal microscopy after 16h at room temperature under dark conditions. The promoter region of TaSRP was amplified by PCR using specific primers. Cis-elements responding to abiotic stresses were analyzed by referring to the plant cis-element database PLACE (http: //www.dna.affrc.go.jp/PLACE). Full length TaSRP cDNA was ligated into the vector pBI121 under control of the CaMV 35S promoter to construct an expression vectors for wild-type Arabidopsis plants. The pBI121-TaSRP construct was introduced into Agrobacterium tumefaciens C58C1 strain cells. Transgenic plants were used for salt resistant experiment. 【Result】 TaSRP, a ricin B-like lectin gene, was obtained by searching the results of salt treatment wheat de novo transcriptome assembly. TaSRP, containing a EUL domain, belongs to EUL family. Homology analysis and multiple alignments showed that there was a high degree of sequence similarity between TaSRP and OSR40 (OSR40g3, OSR40g2 and OSR40c1), but a low degree of sequence similarity between TaSRP and Arabidopsis and G. max. The subcellular localization assay indicated that TaSRP located in cytoplasm. RT-PCR analysis revealed that the expression of TaSRP was upregulated by ABA and salt, but not affected by drought. Isolation of the TaSRP promoter revealed a core promoter element and some cis-acting elements responding to abiotic stresses. Analysis of real-time quantitative PCR showed that TaSRP was up-regulated by high-salt and ABA stresses. The total root lengths of three TaSRP lines were significantly longer than those of wild-type plants in salt treatments (125 mmol•L-1 and 150 mmol•L-1 NaCl) in Arabidopsis plants. These results indicated that TaSRP lines improved salt tolerance of Arabidopsis. 【Conclusion】 TaSRP increased tolerance to salt stress of Arabidopsis plants.

Key words: wheat (Triticum aestivum L.) , EUL family , stress responses , real-time PCR , salt tolerance

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