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High frequency genetic transformation of Cichorium intybus L. using nptII gene as a selective marker

Matvieieva N., Shachovsky A., Kvasko O., Kuchuk N.

 




Cichorium intybus L. is an important vegetable crop used as salad (leaf form) and for the production of coffee substitutes (root form). At the same time these plants can also be used in biotechnologies for synthesis of pharmaceutical proteins. Here we report the possibility of high frequency Agrobacterium rhizogenes- or A. tumefaciens-mediated transformation of C. intybus L. for construction of transgenic hairy roots and plants. The used plasmids contained target human interferon ifn-α2b gene, Mycobacterium tuberculosis ESAT6:Ag85B antigene esxA::fbpBΔTMD fused gene and human telomerase reverse transcriptase hTert gene. Using of nptII gene as a selective one was preferable to the bar gene for chicory. In this case the frequency of transgenic plants or hairy roots formation was significantly higher. Cultivation of explants on the medium with Basta in concentration 12 mg/l have led to plants death or to significant reduction of number of shoots formed. Frequency of hairy roots formation varied from 5,9 to 42,3 % after A. rhizogenes-mediated transformation. Frequency of regeneration of transgenic plants varied from 10 to 86 % after A. tumefaciens-mediated transformation. Both A. rhizogenes- and A. tumefaciens-mediated transformation frequency depended on the type of explants, roots or cotyledons, and vector used. Usage of A. tumefaciens carrying pCB064 plasmid (target esxA::fbpBΔTMD fused gene and nptII selective gene) resulted in the most effective regeneration of transgenic plants with regeneration frequency up to 86 %. In the case of chicory A. rhizogenes-mediated transformation the highest regeneration frequency up to 42,3 % was demonstrated using pCB161 vector with ifn-α2b target gene and nptII selective gene.

Key words: Cichorium intybus L., Agrobacterium-mediated transformation, nptII, bar, human ifn-α2b gene, esxA::fbpBΔTMD gene

Tsitologiya i Genetika 2015, vol. 49, no. 4, pp. 11-16

Institute of Cell Biology and Genetic Engineering, NAS of Ukraine, Kyiv

E-mail: joyna56 gmail.com

Matvieieva N., Shachovsky A., Kvasko O., Kuchuk N. High frequency genetic transformation of Cichorium intybus L. using nptII gene as a selective marker, Tsitol Genet., 2015, vol. 49, no. 4, pp. 11-16.

In "Cytology and Genetics":
N. Matvieieva, A. Shakhovsky, O. Kvasko, N. Kuchuk High frequency genetic transformation of Cichorium intybus L. using nptll gene as a selective marker, Cytol Genet., 2015, vol. 49, no. 4, pp. 220225
DOI: 10.3103/S0095452715040052


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