Citrus (Citrus L.) is the most extensively grown fruit crop worldwide. And there are many in vitro protocols such as callus and cell suspension cultures, organogenesis induction, and protoplast isolation that are viable for genetic transformation. Transgenic citrus plants have been obtained by direct DNA transfer into protoplasts, co-cultivation of internodes or epicotyl segments with Agrobacterium and particle bombardment of nucellar embryogenic cell suspensions. The most widely used method of gene transfer in citrus is the Agrobacterium-mediated transformation of epicotyl segments. Using this system, transgenic plants of citrus species and relatives were obtained, including sweet oranges, sour oranges, limes, grapefruit, and Carrizo citrange rootstock. As for the transgenes, more and more agronomic genes were introduced into citrus plants. These include genes encoding the citrus tristeza virus (CTV) coat protein from different citrus tristeza virus strains (Chap. 9), citrus mosaic virus (CiMV) coat protein, the halotolerance gene HAL2 originally isolated from yeast that confers tolerance to salinity, Arabidopsis LEAFY and APETALA1 genes and citrus FT gene that promote early flower initiation, carotenoid biosynthetic genes from fruits of C. paradisi cv. Flame that control fruit and juice color, and CS-ACS1 gene from Citrus sinensis that controls the ethylene biosynthesis, Xa21 providing broad spectrum Xanthomonas resistance in rice with potential citrus canker disease resistance. And recently, chimaeric ribonuclease gene (barnase) for seedlessness and the juice quality related pectin methylesterase gene (PME) from Valencia orange was introduced into sweet orange by Li et al. (2003) and Guo et al. (2005) respectively.
Among the transformation methods, there are three worthy of note. First, mature tissue transformation. The reliable method for the production of mature transgenic citrus plants via Agrobacterium was proceeded with C. sinen-sis L. Osbeck cultivars 'Pineapple', 'Hamlin', 'Pera', 'Valencia' and 'Natal', though only the former two cultivars succeeded. Second, thin epicotyl sections transformation for Carrizo citrange (C. x Poncirus trifoliata), and Swingle citrumelo (C. paradisi x P. trifoliata) was provided. Third, a further increase in the genetic transformation efficiency in citrus had been obtained by favoring the contact of bacteria and the cambial region. The use of longitudinally cut internodes or epicotyls to expose the cambial region could improve the transformation rate.
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