Obtaining of Transgenic Lettuce Plants, Containing the Recombinant Gene of Thaumatin II Sweet Protein

Ірина Петрівна Копотун, Наталія Леонідівна Щербак

Abstract


Background. At the present, the main direction of genetic engineering research is improving qualities of the crops. The transgenic crops expressing thauII gene successfully combined both the better taste and increasing resistance to fungal pathogens.

Objective. In this study, the recombinant gene of sweet-tasting protein thaumatin II driven by the CaMV 35S promoter was introduced into lettuce (Lactuca sativa) plants.

Methods. The transformation of cotyledonary explants was achieved via Agrobacterium tumefaciens GV3101. The plasmid vectors used in gene integration experiments also contained selective genes of neomycin phosphotransferase II (nptII ) or phosphinothricin acetyl transferase (bar).

Results. The influence of 0,05 % polyvinylpyrrolidone (10000) on the selective concentration of kanamycin and phosphinotricin in the culture medium were investigated. The optimal concentrations of selective agents for stable transgenic plants selection were used.

Conclusions. Availability of the thaumatin II as well as selective nptII or bar genes was verified by the polymerase chain reaction (PCR). An organoleptic analysis of transgenic lettuce plants did not reveal significant changes in the taste quality. The leaf extracts of these plants will be further tested for antifungal activity.

 


Keywords


Transgenic lettuce; Thaumatin II; Polyvinylpyrrolidone; Kanamycin; Phosphinothricin

References


T. Masuda and N. Kitabatake, “Developments in biotechnological production of sweet proteins”, J. Biosci. Bioeng., vol. 102, no. 5, рр. 375–389, 2006.

L. Edens et al., “Cloning of cDNA encoding the sweet-tasting plant protein thaumatin and its expression in Eschericha coli”, Gene, vol. 18, рр. 1–12, 1982.

Ch. Illingworth et al., “Secretion of the sweet-tasting plant protein thaumatin by Streptomyces lividans”, J. Industrial Microbiol., vol. 4, pp. 37–42, 1989.

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A.V. Kuz’menko et al., “Creating transgenic lettuce plants containing cross-linked gene protein antigens Esat6:Ag85b with Mycobacterium tuberculosis”, Naukovi Visti NTUU KPI, no. 3, pp. 65–69, 2013 (in Ukrainian).

O. Gamborg et al., “Nutrient reguirements of suspesion cultures of soybean root cells”, Exp. Cell Res., vol. 50, pp. 151–158, 1968.

J.J. Doyle and J.L. Doyle, “Isolation of plant DNA from fresh tissue”, Focus, vol. 12, pp. 13–15, 1990.

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GOST Style Citations


  1. Masuda T., Kitabatake N. Developments in biotechnological production of sweet proteins // J. Biosci. Bioeng. – 2006. – 102, № 5. – P. 375–389. 
       
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  3. Illingworth Ch., Larson G., Hellekant G. Secretion of the sweet-tasting plant protein thaumatin by Streptomyces lividans // J. Industrial Microbiol. – 1989. – 4. – P. 37–42. 
       
  4. Pham N.B., Schäfer H., Wink M. Production and secretion of recombinant thaumatin in tobacco hairy root cultures // Biotechnol. J. – 2012. – 7. – P. 537–545. 
       
  5. Variable properties of transgenic cucumber plants containing the thaumatin II gene from Thaumatococcus daniellii / M. Szwacka, M. Krzymowska, A. Osuch et al. // Acta Physiologiae Plantarum. – 2002. – 24, № 2. – P. 173–185. 
       
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DOI: https://doi.org/10.20535/1810-0546.2015.3.61497

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