Abstract

VACA-VACA, Juan Carlos; PULIDO-RENDON, Andrea Jimena  and  LOPEZ-LOPEZ, Karina. Optimization of low pressure biobalistics conditions for analysis of transient expression of heterologous gene in tobacco leaves tobacco cultivated in vitro. Acta Agron. [online]. 2015, vol.64, n.2, pp.146-155. ISSN 0120-2812.  https://doi.org/10.15446/acag.v64n2.43928.

Transient expression studies of heterologous genes with biotechnological interest are a prerequisite to its stable expression in transgenic plants. The aim of this research was to optimized the biobalistic conditions for testing transient expression of heterologous genes in tobacco (Nicotiana tabacum - Xanthi) using the low pressure Helios® Gene Gun System (Biorad ®). As heterologous gene construction was used CaMV 35S promoter fused to the uidA (GUS) gene cloned into the vector pBI121. The optimized conditions were: shot pressure, number of shots and shooting distance of leaf fragments or full sheets of tobacco. GUS expression was evaluated by average number of the blue dots observed on plant tissues already bombarded. Only blue dots were observed when whole leaves tobacco were used. A great deal of blue spots was observed using the following conditions: a pressure of 160 psi, 4 shots and 0 cm on whole sheets. In order to confirm the conditions earlier obtained, the pCAMBIA 1305.2 vector was employed in this experiment. Higher numbers of blue points were observed in tissues previously bombarded; those were as an indicative of the efficiency experimental conditions earlier optimized.

Keywords : β Glucuronidase; transient expression; Helios® Gene Gun System; Geminivirus.

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