Improvement of organic phosphate acquisition in transgenic tobacco plants by overexpression of a soybean phytase gene Sphy1

Abstract

Due to the huge amount in the soil, phytate is an important potential source for providing the plants with available phosphorus (Pi) by the involved catalytic reaction of phytase. In this study, a construct fusing the open reading frame (ORF) of Sphy1 into corresponding positions in the fragment of binary expression vector pBI121 was created and used to transform tobacco. Molecular identification by PCR and RT-PCR indicated the target gene Sphy1 in the transgenic tobacco plants was transcribed under the regulation of an upstream promoter. Compared with the control plants, the phytase activities in all the transgenic plants were increased, with the increased range consistent with the expression levels in the transgenic plants. Under the growth conditions with phytate as the sole phosphorus source, the transgenic line 1 plants displayed a high expression level of Sphy1 and shows notable improved growth performance, such as higher fresh weight and dry weight, as well as higher total P content and more accumulative P amount per plant than CK. This clearly indicated that overexpression of Sphy1 could improve the phosphorus acquisition by the extruded Sphy1 phytase in the rhizosphere, where this enzyme could catalyze the degradation of the phytate and release the available Pi for plants. The Sphy1 gene seemed to have a potential value in the creation of new crop cultivars with high phosphorus use efficiency.

Keywords

Sphy1 gene / overexpression / phosphorus utilization / transgenic tobacco

References

[1] Abelson P H (1999). A potential phosphate crisis. Science,283: 2015-2021

[2] Anderson G.(1980). Assessing organic phosphorus in soils. In: Khasawneh F E, Sample E C, Kamprath E J, eds. The Role of Phosphorus in Agriculture. Madison: American Society of Agronomy, 411-431

[3] Bieleski R L (1973). Phosphate pools, phosphate transport, and phosphate availability. Annu Rev Plant Physiol, 24: 225-252

[4] Chen M H, Huang L F, Li H M, Chen Y R, Yu S M (2004). Signal peptide-dependent targeting of a rice α-Amylase and Cargo proteins to plastids and extracellular compartments of plant cells. Plant Physiol, 135: 1367-1377

[5] Dalal R C (1977). Soil organic phosphorus. Adv Agron, 29: 83-117

[6] Guo L, Wang J J, Xiao K (2008). Isolation and characterization of a novel phytase gene (Sphy1) from soybean (Glycine max L.). Frontier of Agriculture in China, 2(2): 30-36

[7] Han S F, Gu J T, Xiao K (2007). Improving organic phosphate utilization in transgenic white clover by overexpression of Aspergillus niger PhyA gene. Frontier of Agriculture in China, 1(3): 265-270

[8] Hood E E, Gelvin S B, Melchers L S, Hoekema A (1993). New Agrobacterium helper plasmids for gene transfer to plants. Transgenic Research, 2(4): 208-218

[9] Kavanová M, Grimoldi A A, Lattanzi F A, Schnyder H (2006). Phosphorus nutrition and mycorrhiza effects on grass leaf growth. P status- and size-mediated effects on growth zone kinematics. Plant Cell Environ, 29(4): 511-520

[10] Li J, Hegeman C E, Hanlon R W, Lacy G H, Denbow D M, Grabau E A (1997). Secretion of active recombinant phytase from soybean cell-suspension cultures. Plant Physiol, 114: 1103-1111

[11] Manfield I W, Pavlov V K, Li J, Cook H, Hummel F, Gilmartin P M (2005). Molecular characterization of DNA sequences from the Primula vulgarisS-locus. J Exp Bot, 56: 1177-1188

[12] Murphy J, Riley J P (1962). A modified single solution method for the determination of phosphate in natural waters. Analytica Chimica Acta, 27: 31-36

[13] Raghothama K G (1999). Phosphate acquisition. Annu Rev Plant Physiol, 50: 665-693

[14] Richardson A E, Hadobas P A, Hayes J E (2001). Extracellular secretion of Aspergillus phytase from Arabidopsis roots enables plants to obtain phosphorus from phytate. Plant J, 25: 641-649

[15] Vance C P, Uhde-Stone C, Allan D L (2003). Phosphorus acquisition and use: critical adaptation by plants for securing a nonrenewable resource. New Phytologist, 157: 423-447

[16] Von Uexku H R, Mutert E (1995). Global extent, development and economic impact of acid soils. Plant Soil, 171: 1-15

[17] Xiao K, Harrison M J, Wang Z Y (2005). Transgenic expression of a novel M. truncatula phytase gene results in improved acquisition of organic phosphorus by Arabidopsis. Planta, 222: 27-36

[18] Zhang H N, Li X J, Li C D, Xiao K (2008). Effects of overexpression of wheat superoxide dismutase (SOD) genes on salt tolerant capability in tobacco. Acta Agronomica Sinica, 34(8): 1403-1408

[19] Zimmermann P, Zardi G, Lehmann M, Zeder C, Amrhein N, Frossard E, Bucher M (2003). Engineering the root-soil interface via targeted expression of a synthetic phytase gene in trichoblasts. Plant Biotechnol, 1: 353-360

Acknowledgements

This work was supported by the Natural Science Foundation of Hebei Province, China (No. C2006000434), the Ph.D. Programs Foundation of Ministry of Education of China (No. 20060086001)and the National Basic Research Program of China (973) in the early stage (No.2007CB116209).

RIGHTS & PERMISSIONS

2014 Higher Education Press and Springer-Verlag Berlin Heidelberg