Bao-Tai Guo, Bin Wang, Man-Li Weng, Li-Xian Qiao, Yan-Bin Feng, Li Wang, Peng-Yan Zhang, Xiu-Liang Wang, Jiong-Ming Sui, Tao Liu, De-Lin Duan, Bin Wang


The trehalose-6-phosphate synthase gene of Porphyra yezoensis (PyTPS) was isolated and cloned into a plant gene expression vector pCAMBIA2300-35S-OCS, and the resulting construct pCAMBIA2300-PyTPS was transformed into Agrobacterium tumefaciens (A. tumefaciens ) strain AGL1. Genetic transformation of rice variety TP309 was performed with the A. tumefaciens containing pCAMBIA2300-PyTPS. After antibiotic G418 screening and PCR analysis, one hundred T0 transgenic plants were seclected and transplanted into the trial field in the greenhouse and used for further study. Ninety-five of these 100 T0 transgenic cultivaries produced their seeds, which were harvested and stored separately. All of the 95 potential T1 transgenic lines were re-identified by PCR analysis, and their salt-tolerance was tested with 3‰ and 5‰ NaCl solutions. Results indicated that 78 of the 95 T1 transgenic lines were PCR- positive and resistant to 5‰ NaCl solution. Salt-tolerance of these 78 T1 transgenic lines was further tested with higher concentration of NaCl solutions. Of which, three lines (H155, H191 and Y308) showed resistance to 8‰ NaCl in the test. These 3 lines were comprehensively analyzed by PCR, Southern hybridization, northern hybridization and RT-PCR analyses. In addition, trehalose content measurement and preliminary yield evaluation were carried out, results indicated that the PyTPS gene was integrated into the genomic DNA sequences of these 3 transgenic lines and expressed indeed in the transgenic plants. Detection of the transformed PyTPS gene in these 3 transgenic lines was performed in plants from T1 to T6 generations; results indicated that the transformed PyTPS gene was present in transgenic plants from T1 to T6 generations.


Agrobacterium-mediated transformation, Porphyra yezoensis Salt-tolerance, Transgenic rice, Trehalose-6- phosphate synthase (TPS)

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Altman, A., Hasegawa, M.H., 2012. Plant Biotechnology and Agriculture. XXVii. Academic Press, San Diego, USA.

Avonce, N., Leyman, B., Mascaras-Gallardo, J.O., van Dijck, P., Thevelein, J.M., Iturriaga, G., 2004. The Arabidopsis trehalose-6-P-synthase AtTPS1 gene is a regulator of glucose, abscisic acid, and stress signaling. Plant Physiol. 136, 3649–3659.

Chomczynski, P., Sacchi, N., 1987. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal. Biochem. 162, 156–159.

Eastmond, P.J., van Dijken, A.J., Spielman, M., Kerr, A., Tissier, A.F., Dickinson, H.G., Jones, J.D., Smeekens, S.C., Graham, I.A., 2002. Trehalose-6-phosphate synthase 1, which catalyses the first step in trehalose synthesis, is essential for Arabidopsis embryo maturation. Plant J. 29, 225–235.

Elbein, A.D., 1974. The metabolism of alpha, alpha-trehalose. Adv. Carbohydr. Chem. Biochem. 1974, 30, 227–256.

Garg, A.K., Kim, J.K., Owens, T.G., Ranwala, A.P., Choi, Y.D., Kochian, L.V., Wu, R.J., 2002. Trehalose accumulation in rice plants confers high tolerance levels to different abiotic stress. Proc. Natl. Acad. Sci. USA. 99, 15898–15903.

Goddijn, O.J.M., Verwoerd, T.C., Voogd, E., Krutwagen, R.W.H.H., de Graaf, P.T.H.M., Poels, J., van Dun, K., Ponstein, A.S., Damm, B., Pen, J., 1997. Inhibition of trehalase activity enhances trehalose accumulation in transgenic plants. Plant Physiol. 113, 181–190.

Goddijn, O.J.M., Smeekens, S., 1998. Sensing trehalose biosynthesis in plants. Plant J. 14, 143–146.

Goddijn, O.J.M., van Dun, K., 1999. Trehalose metabolism in plants. Trends Plant Sci. 1999, 4, 315–319.

Hiei, Y., Ohta, S., Komari, T., Kumashiro, T., 1994. Efficient transformation of rice (Oryza sativa L.) mediated by Agrobacterium and sequence analysis of the boundaries of the T-DNA. Plant J. 6, 271–282.

Holmstrom, K.O., Mantyla, E., Welin, B., Manadal, A., Palva, E.T., Tunnela, O.E., Londesborough, J., 1996. Drought tolerance in tobacco. Nature, 379, 683–684.

Jang, I.C., Oh, S.J., Seo, J.S., Choi, W.B., Song, S.I., Kim, C.H., Kim, Y.S., Seo, H.S., Choi, Y.D., Nahm, B.H., Kim, J.K., 2003. Expression of bifunctional fusion of the Escherichia coli genes for trehalose-6-phosphate synthase and trehalose-6-phosphate phosphatase in transgenic rice plants increases trehalose accumulation and abiotic stress tolerance without stunting growth. Plant Physiol. 131, 516–524.

Jia, W.L., Hu, Y.L., Zhang, Y.Q., Yang, L.L., Lin, Z.P., Wu, Q., 2007. Transformation of trehalose synthase gene (TPS gene) into perennial ryegrass and identification of drought tolerance. Mol. Plant Breed. 5, 27–31 (In Chinese with English abstract).

Jia, J., Cui, X.J., Wu, J.H., Wang, J.Y., Wang, G.Z., 2011. Physiological and biochemical responses of halophyte. Afr J Biotechnol, 10, 11468–11476.

Jeong, S.C., Pack, I.S., Cho, E.Y., Youk, E.S., Park, S., Yoon, W.K., Kim, C.G., Choi, Y.D., Kim, J.K., Kim, H.M., 2007. Molecular analysis and quantitative detection of a transgenic rice line expressing a bifunctional fusion TPSP. Food Control, 18, 1434–1442.

Kaasen, I., McDougall, J., Strom, A.R., 1994. Analysis of the otsBA operon for osmoregulatory trehalose synthesis in Escherichia coli and homology of the OtsA and OtsB proteins to the yeast trehalose-6-phosphate synthasephosphatase complex. Gene, 145, 9–15.

Karim, S., Aronsson, H., Ericson, H., Pirhonen, M., Leyman, B., Welin, B., Mantyla, E., Palva, E.T., van Dijck, P., Holmstrom, K.O., 2007. Improved drought tolerance without undesired side effects in transgenic plants producing trehalose. Plant Mol. Biol. 64, 371–386.

Kosmas, S.A., Argyrokastritis, A., Loukas, M.G., Eliopoulos, E., Tsakas, S., Kaltsikes, P.J., 2006. Isolation and characterization of drought-related trehalose-6-phosphate synthase gene from cultivated cotton (Gossypium hirsutum L.). Planta, 223, 329–339.

Liu, J., Xiang, D.Y., Chen, J.B., Zhan, W., 2009. Low temperature LT50 of three Eucalyptus seedlings with electrical conductivity method and logistic equation (in Chinese). Guangxi Forestry Science, 38, 75–79 (in Chinese with English abstract).

Lowe, R.G., Lord, M., Rybak, K., Trengove, R.D., Oliver, R.P., Solomon, P.S., 2009. Trehalose biosynthesis is involved in sporulation of Stagonospora nodorum. Fungal Genet. Biol. 46, 381–389.

Lunn, J.E., Feil, R., Hendriks, J.H.M., Gibon, Y., Morcuende, R., Osuna, D., Scheible, W.R., Carillo, P., Hajirezaei, M.R., Stitt, M., 2006. Sugar-induced increases in trehalose 6-phosphate are correlated with redox activation of ADP-glucose pyrophosphorylase and higher rates of starch synthesis in Arabidopsis thaliana. Biochem. J. 397, 139–148.

Mu, Y., He, J., Fu, F.L., Li, W.C., 2007, Transformation of maize with trehalose synthase gene (TPS1) cloning from Saccharomyces cerevisiae. J. Nucl. Agric. Sci. 21, 430–435 (In Chinese with English abstract).

Muller, J., Wiemken, A., Aeschbacher, R., 1999. Trehalose metabolism in sugar sensing and plant development. Plant Sci. 147, 37–47.

Paul, M.J., Primavesi, L.F., Jhurreea, D., Zhang, Y.H., 2008. Trehalose metabolism and signaling. Ann. Rev. Plant Biol. 59, 417–441.

Pilon-Smits, E.A.H., Terry, N., Sears, T., Kim, H., Zayed, A., Hwang, S., van Dun, K., Voogd, E., Verwoerd, T.C., Krutwagen, R.W.H.H., 1998. Trehalose-producing transgenic tobacco plants show improved growth performance under drought stress. J. Plant Physiol. 152, 525–532.

Romero, C., Belles, J.M., Vaya, J.L., Serrano, R., Culianez-Macia, F.A., 1997. Expression of the yeast trehalose-6-phosphate synthase gene in transgenic tobacco plants: pleiotropic phenotypes include drought tolerance. Planta, 201, 293–297.

Sambrook, J., Fritsch, E.F., Maniatis, T., 1989. Molecular Cloning: A Laboratory Manual, 2nd ed.; Cold Spring Harbor Laboratory Press: Cold Spring Harbor, NY, USA, pp 7.37-7.52, pp 9.31–9.58.

Sukumaran, N.P., Weiser, C.J., 1972. An excised leaflet test for evaluating potato frost tolerance. HortScience, 7, 467–468.

Vogel, G.; Fiehn, O., Jean-Richaro-dit-Bresse, L., Boller, T., Wiemken, A., Aeschbacher, R.A., Wingler, A., 2001. Trehalose metabolism Arabidopsis: occurrence of trehalose and molecular cloning and characterization of trehalose-6-phosphate synthase homologs. J. Exp. Bot. 52, 1817–1826.

Vuorio, O.E., Kalkkinen, N., Londesborough, J., 1993. Cloning of two related genes encoding the 56-kDa and 123-kDa subunits of trehalose synthase from the yeast Saccharomyces cerevisiae. Eur. J. Biochem. 216, 849–861.

Wang, G.L., Zhao, G., Feng, Y.B., Xuan, J.S., Sun, J.W., Guo, B.T., Jiang, G.Y., Weng, M.L., Yao, J.T., Wang, B., Duan, D.L., Liu, T., 2010. Cloning and comparative studies of seaweed trehalose-6-phosphate synthase genes. Mar. Drugs, 8, 2065–2079.

Wang, Y.G., Xing, Q.H., Deng, Q.Y., Liang, F.S., Yuan, L.P., Weng, M.L., Wang, B., 2003. Fine mapping of the rice thermo-sensitive genic male-sterile gene tms5. Theor. Appl. Genet. 107, 917–921.

Wu, L.Q., Fan, Z.M., Guo, L., Li, Y.Q., Zhang, W.J., Qu, L.J., Chen, Z.L., 2003. Over-expression of an Arabidopsis δ-OAT gene enhances salt and drought tolerance in transgenic rice. Chinese Science Bulletin, 48, 2594-2600.

Zhao, F., Li, Q.Y., Weng, M.L., Wang, X.L., Guo, B.T., Wang, L., Wang, W., Duan, D.L., Wang, B., 2013. Cloning of TPS gene from eelgrass species Zostera marina and its functional identification by genetic transformation in rice. Gene, 531, 205–211.


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