Next generation synthetic vectors for transformation of the plastid genome of higher plants
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Plastid transformation vectors are E. coli plasmids carrying a plastid marker gene for selection, adjacent cloning sites and flanking plastid DNA to target insertions in the plastid genome by homologous recombination. We report here on a family of next generation plastid vectors carrying synthetic DNA vector arms targeting insertions in the rbcL-accD intergenic region of the tobacco (Nicotiana tabacum) plastid genome. The pSS22 plasmid carries only synthetic vector arms from which the undesirable restriction sites have been removed by point mutations. The pSS24 vector carries a c-Myc tagged spectinomycin resistance (aadA) marker gene whereas in vector pSS30 aadA is flanked with loxP sequences for post-transformation marker excision. The synthetic vectors will enable direct manipulation of passenger genes in the transformation vector targeting insertions in the rbcL-accD intergenic region that contains many commonly used restriction sites.
KeywordsHomologous recombination Nicotiana tabacum Plastid transformation Tobacco Vector
Sugey Ramona Sinagawa-García was supported by a Ph.D. scholarship from Consejo Nacional de Ciencia y Tecnología (CONACYT), México.
- Horvath EM, Peter SO, Joët T, Rumeau D, Cournac L, Horvath GV, Kavanagh TA, Schäfer C, Peltier G, Medgyesy P (2000) Target inactivation of the plastid ndhB gene in tobacco results in an enhanced sensitivity of photosynthesis to moderate stomatal closure. Plant Physiol 123:1337–1350. doi: 10.1104/pp.123.4.1337 PubMedCrossRefGoogle Scholar
- Koop HU, Herz S, Golds TJ, Nickelsen J (2007) The genetic transformation of plastids. In: Bock R (ed) Cell and molecular biology of plastids (topics in current genetics). Springer Verlag, Berlin, pp 457–510Google Scholar
- Shinozaki K, Ohme M, Tanaka M, Wakasugi T, Hayashida N, Matsubayashi T, Zaita N, Chunwongse J, Obokata J, Yamaguchi-Shinozaki K, Ohto C, Torazawa K, Meng B-Y, Sugita M, Deno H, Kamogashira T, Yamada K, Kusuda J, Takaiwa F, Kato A, Tohdoh N, Shimada H, Sugiura M (1986) The complete nucleotide sequence of the tobacco chloroplast genome: its gene organization and expression. EMBO J 5:2043–2049PubMedGoogle Scholar
- Yukawa M, Tsudzuki T, Sugiura M (2006) The chloroplast genome of Nicotiana sylvestris and Nicotiana tomentosiformis: complete sequencing confirms that the Nicotiana sylvestris progenitor is the maternal genome donor of Nicotiana tabacum. Mol Genet Genomics 275:367–373. doi: 10.1007/s00438-005-0092-6 PubMedCrossRefGoogle Scholar
- Zhou F, Badillo-Corona JA, Karcher D, Gonzalez-Rabade N, Piepenburg K, Borchers AM, Maloney AP, Kavanagh TA, Gray JC, Bock R (2008) High-level expression of human immunodeficiency virus antigens from the tobacco and tomato plastid genomes. Plant Biotechnol J 6:897–913. doi: 10.1111/j.1467-7652.2008.00356.x CrossRefGoogle Scholar