A protocol for Agrobacterium-mediated transformation in rice

Abstract

Agrobacterium-mediated transformation of rice is an important method that has been widely adopted by many laboratories. However, because current approaches rely on culture systems, routine protocols have been established only in japonica rice, especially those varieties with higher regeneration potential. Some very efficient methods have been developed for japonica varieties that enable high-throughput functional analysis in rice; however, many elite japonica, and most indica, varieties are difficult to regenerate, leading to low transformation efficiencies. Much effort has been devoted to improving transformation efficiency for all rice genotypes. Here, we describe an Agrobacterium-mediated rice transformation method that is applicable to easily cultured varieties in addition to elite japonica varieties that are more difficult to culture. Using this method, transgenic rice plants can be obtained in about 2-3 months with a transformation frequency of 30-50%, both in easily cultured varieties and recalcitrant elite japonica rice.

Genetic engineering of gene encoding transcription factor MtOsDREB2A form Vietnamese rice variety Chanh trui for drought stress tolerance

“…KÝch th−íc 2,9 kb t−¬ng øng víi sù céng dån kÝch th−íc gien MtOsDREB2A MtOsDREB2A vµo gièng lóa Chµnh trôi th«ng qua Agrobacterium EHA105. Toµn bé qu¸ tr×nh h×nh thµnh callus, l©y nhiÔm vµ t¸i sinh c©y ®−îc tiÕn hµnh theo c¸c ph−¬ng ph¸p ® ®−îc tèi −u bëi Nishimura [5]. Tõ 100 h¹t lóa gièng Chµnh trôi ®−îc vµo nu«i cÊy, chóng t«i thu ®−îc 91 callus sau 10 ngµy nu«i cÊy trªn m«i tr−êng MS cã bae sung 2 mg/l 2,4D.…”

“…ViÖc ¸p dông thµnh c«ng kü thuËt nµy cho phÐp chóng t«i cã thÓ më réng vµ tiÕn hµnh ®ång thêi c¸c thÝ nghiÖm chuyÓn gien kh¸c nhau vµo c©y lóa. Quy tr×nh chuyÓn gien th«ng qua vi khuÈn A.tumefaciens cña nhãm t¸c gi ¶ Nishimura [5] mAEc dï ®−îc tèi −u trªn c©y lóa Japonica nh−ng vÉn thÝch hîp víi c©y lóa Chµnh trôi (gièng indica) cña ViÖt Nam víi hiÖu suÊt chuyÓn gien ®¹t tíi 25%. C¸c c©y chuyÓn gien sau ®ã cã kh ¶ n¨ng sinh tr−ëng b×nh th−êng trong ®iÒu kiÖn nhµ kÝnh còng nh− trªn ruéng thÝ nghiÖm.…”

Bamboo Regeneration via Embryogenesis and Organogenesis

“…The NiR gene cloned from the high-regeneration rice strain Konansou has the similar function with that of Kasalath (Ozawa & Kawahigashi, 2006). However, NiR gene based improvement method will be suitable for the major Japonica rice varieties but not Indica rice varieties for having high NiR activity (Nishimura et al, 2006). In addition, the NiR gene can be used as a selection marker for rice gene transformation (Nishimura et al, 2005;Ozawa & Kawahigashi, 2006).…”

Differential transformation efficiency of Japonicarice varieties developed in northern China

“…Agrobacterium-mediated transformation is a general method for genetic modification in many plant species because it allows efficient insertion of stable, unrearranged, single-copy sequences into plant genomes (Nishimura et al 2006). Efficient methods for Agrobacterium-mediated transformation of japonica rice have been developed (Hiei et al 1994, Hiei et al 1997, Hiei and Komari 2008.…”

“…Two japonica varieties, Nipponbare and Kitaake (developed in Japan), are good genotypes for Agrobacterium-mediated transformation. Based on this method, Agrobacteriummediated transformation methods have been improved, with minor modification for many rice varieties, not only japonica but also indica and tropical japonica (Nishimura et al 2006, Hiei and Komari 2008, Saika and Toki 2010.…”