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Christiansen, P., Andersen, C.H., Didion, T., Folling, M., Nielsen, K.K. (2005) A rapid and efficient transformation protocol for the grass Brachypodium distachyon. Plant Cell Reports, 23,751-758.

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• TITLE: Rapid regeneration of stable transformants in cultures of potato by improving factors influencing Agrobacterium-mediated transformation

  AUTHORS: Bipasha Chakravarty, Gefu Wang-Pruski

  KEYWORDS: Bacterial Concentration; Cultivar; Explant; Potato Transformation; Preculture

  JOURNAL NAME: Advances in Bioscience and Biotechnology, Vol.1 No.5, December 30, 2010

  ABSTRACT: An efficient and rapid Agrobacterium tumefaciens-mediated transformation protocol was developed to generate activation-tagged mutant lines with the aim of large-scale functional analysis of the potato genome. The explants were inoculated with an Agrobacterium strain harboring the binary plasmid pSKI074 containing four CaMV 35S enhancers in the T-DNA region which activates the downstream genes in the host plant after its integration. Various parameters investigated to increase transformation efficiency were the type and age of explant, cultivar, hormone combinations, preculture of explants, period of co-cultivation with bacteria and concentration of bacterial cultures used for transformation. Stem explants from 5 week old plantlets of cv. Bintje which had undergone phytohormone pretreatment for 4 days, inoculation with diluted bacterial concentration of OD600 = 0.2 containing acetosyringone followed by 2 days of co-cultivation and selection in media with IAA and trans-zeatin all helped in greatly improving the transformation efficiency. The total time required from infection to rooted shoots was 6-7 weeks. Initial evidence for stable integration and expression of the transgenes by PCR analysis showed that over 93% of the regenerated lines were transgenic and this was confirmed by Southern hybridization.