CBN-V Video Archives - S5-03
Isolation and Characterization of Three Promoters Directing Gus-expression to Cassava Roots

Bohl Zenger S., P. Zhang, J. Puonti-Kaerlas, I. Potrykus and W. Gruissem

Institute for Plant Science, ETH-Zentrum/LFW E17, CH-8092, Zürich zhang.peng@ipw.biol.ethz.ch

        Presently, there are few tissue specific promoters available for targeted engineering of cassava. Root specific promoters are especially lacking. In order to isolate tissue specific cDNAs, we differentially screened a tuber cDNA library and were able to isolate a number of potentially root specific cDNAs. Northern blot and RT-PCR confirmed the specificity of the isolated sequences. A genomic library was constructed and screened, with some selected cDNAs and corresponding genes and promoters isolated. Promoters p15 (1.5 kb) and p54 (1 kb) were identified, sequenced and fused to the uidA (Gus) gene in order to analyze their activity in transgenic Arabidopsis and cassava. A third promoter-Gus fusion gen (1.1 kb)-Gus was provided by Marcel Bucher, ETH Zurich. gen is a root hair cell specific promoter isolated from tomato. In Arabidopsis seedlings p15 and p54 were both mainly active in vascular tissue, gen in the entire root. In transgenic cassava in vitro roots p15-Gus showed expression in different cell types of the central cylinder, while p54-Gus showed expression in cortex cells and the central cylinder. gen-Gus was exclusively active in the rhizodermis of in vitro roots. Since it is known from RT-PCR that the isolated promoters p15 and p54 have altered specificity under in vitro conditions, transgenic plantlets will soon be transferred to soil to prove the specificity of all the promoters under greenhouse conditions.

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