New Ac/Ds-based constructs for efficient gene and enhancer trapping in rice

摘要

We have evaluated Ac/Ds gene and enhancer trapping constructs and shown that all components except the negative selector nptll gene work in rice. Hygromycin counter selection could identify the possible stable trap lines that did not contain Ac and inwhich Ds had jumped to locations unlinked to their original positions, However, high-throughput screening requires quick and efficient selection of stable Ds trap lines, linked or unlinked to their original positions, from among the screening population. We have evaluated two negative selectors, tms_2 (JaaH) under the control of the manopine synthase 2 promoter from Agrobacterium tumefaciens and codA driven by the CaMV 358 promoter, for use in rice. The former was found to be suitable for rice and an Ac construct with tms_2 as a negative selector has been employed. In the Ds enhancer and gene trap constructs, the nptll gene has been replaced with bar as a strong negative selector for Ds. Ds gene and enhancer trap constructs with bar as a Ds excision marker have also been made, Use of these new constructs will simplify the screening for stable trap lines.