Stable Transformation of Embryogenic Tissues of Pinus nigra Arn. using a Biolistic Method
Slovak Academy of Sciences, Presburg, Bratislavský, Slovakia Biotechnology Letters
(Impact Factor: 1.59).
08/2005; 27(13):899-903. DOI: 10.1007/s10529-005-7178-4
The stable transformation of embryogenic tissues of Pinus nigra Arn., cell line E104, has been achieved using a biolistic approach. The introduced DNA consisted of the uidA reporter gene under the control of the double CaMV 35S promoter and the nptII selection gene controlled by the single CaMV 35S promoter. Three days after bombardment, putative transformed tissues were selected for continued proliferation on a medium containing 20 mg geneticin l(-1). Resistant embryogenic tissue recovery required 10-12 weeks. The integration of the nptII and uidA genes was confirmed by both histochemical/fluorimetric GUS assays and PCR amplification of the inserts in the five geneticin resistant sub-lines of line E104. The activity of the uidA reporter gene in transgenic, embryogenic tissue lines was stable and could be detected after one year of culture. Somatic embryo maturation was, however, poor and no plantlet regeneration could be obtained.
Available from: Krystyna Klimaszewska
- "AT, LISW, PB S Stomp et al. 1990; Newton et al. 1993; Tang et al. 2006b griffithii PB T Fernando et al. 2000 halepensis AR S Tzfira et al. 1996 jeffreyi AT Gall Stomp et al. 1990 lambertiana AT S Loopstra et al. 1990 maximartinezii AT S Villalobos-Amador et al. 2002 monticola AR, PB T McAfee et al. 1993; Fernando et al. 2000 nigra AR, AT, EP, PB S Mihaljevic et al. 1996; Lopez et al. 2000a; Salaj et al. 2005 palustris AR, AT, PB T Diner 1999 patula PB SP Nigro et al. 2004 pinaster AT, EP, PB SP Gomez-Maldonado et al. 2001; Trontin et al. 2002 pinceana AT S Villalobos-Amador et al. 2002 pinea AT, PB, SAAT T Humara et al. 1999a, 1999b ponderosa AT Gall Morris et al. 1989; Stomp et al. 1990 radiata AT, EP, PB SP Stomp et al. 1990; Campbell et al. 1992; Walter et al. 1998; Charity et al. 2005 rigida x taeda AT, PB SP Connett-Porceddu et al. 2003, 2007 roxburghii PB SP Parasharami et al. 2006 strobus AT, PB SP Tian et al. 1997; Levée et al. 1999 sylvestris AT, AR, PB SP Stomp et al. 1990; Aronen et al. 2003 taeda AT, PB, SAAT SP Sederoff et al. 1986; Tang et al. 2001; Connett-Porceddu et al. 2003, 2007 taeda x elliottii AT Gall Huang and Tauer 1994 thunbergii PB T Taniguchi et al. 2004 virginiana AT, PB SP Stomp et al. 1990; Tang and Newton 2004b, 2005 the genome by a variety of recombination events. In the AT method, the gene(s) of interest are inserted into the transfer- DNA (T-DNA) region of tumor-inducing plasmid (pTi) from a disarmed A. tumefaciens strain. "
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ABSTRACT: Advances in conifer tissue culture and genetic transformation offer new opportunities in the field of genetic engineering. Genetic
transformation is an important tool for breeders because it allows them to introduce valuable genes that might otherwise be difficult to
integrate into elite genotypes. Combined with mass vegetative propagation, such as somatic embryogenesis, regeneration of transgenic
conifers could result in accelerated tree improvement. Genetic transformation requires the development of an efficient gene delivery
system. Extensive work has been done in Pinus spp. to improve transformation methods, either by DNA-coated particle bombardment or
by co-culture with Agrobacterium tumefaciens. This review describes the most recent progress made in genetic transformation in the
genus Pinus, with emphasis on four important forest and plantation species grown in Europe (P. pinaster), New Zealand, Australia and
South America (P. radiata), and North America (P. taeda and P. strobus). The biosafety issues associated with potential deployment of
transgenic pine varieties in commercial forestry are highlighted.
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ABSTRACT: In this review we examine the history and progression of conifer genetic engineering. The review includes the methods used, the conifer species transformed, the genes inserted and the regeneration of genetically engineered conifer trees. We cover both Biolistic® and Agrobacterium-mediated transformation, and we detail transformation events with and without plant regeneration. We show that almost all conifer transformation work uses nptII as a selective marker, and very often uidA is included as a reporter gene. Further, we show that a range of genes that are of commercial interest for forest tree plantations have been introduced, such as herbicide resistance, insect resistance and those related to wood properties. We briefly discuss the future for biotechnology in the context of socially acceptable enhanced plantation forestry and under consideration of benefits and risks.
Available from: Elsbeth Lewis Walker
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ABSTRACT: In developing and developed nations, plant cell culture systems are used to supply desirable compounds in lieu of chemical synthesis or natural extraction. When plant cell culture systems are unable to meet commercial demand, metabolic engineering offers a method to increase yields. However, to benefit from metabolic engineering approaches, effective transient transformation methods are required to rapidly identify and characterize key regulatory genes before intensive, time-consuming stable transformation efforts can proceed. This paper describes a particle bombardment-mediated transient transformation system for Taxus spp. in cell culture. Optimal parameters were established for the T. cuspidata cell line P991 and the T. canadensis cell line CO93D, resulting in reliable, efficient, transient expression of the firefly luciferase gene under control of the constitutive CaMV 35S promoter. Multiple bombardments and larger gold microcarriers (1.6 vs 1.0 microm in diameter) were particularly effective in increasing luciferase activity and in reducing variation among replicates. This particle bombardment-mediated transformation system was also shown to be capable of transiently expressing the DsRed and beta-glucuronidase reporter genes under the control of the maize ubiquitin and CaMV 35S promoters, respectively. With the ability to transiently transform Taxus spp. cell cultures using a variety of promoters and reporters, characterization of genes related to paclitaxel accumulation in culture can now proceed.
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