Study Of Transformation And Expression Of Heavy-metals Resistant Genes OASA4and γ-GCS Of Brassica Chinensis In Nicotiana Tabacum

Heavy metal pollutants degrade environmental quality and further hazard humanmetabolism by their accumulation in food chain. Phytoremediation to clean up heavymetal as an environmental friendly and cost effective method has attracted increasingattention in the past decades. Pre-existing wild heavy metal hyperaccumulators haveseveral inherent shortcomings, such as low level of heavy metal accumulation, slow andincomplete process, small biomass, long growth cycle, and so on. Transgenic approachis promising and suitable to overcome the limitations of detoxification capabilities ofwild plant. During the last decade, transgenic plants have been successfully employed topromote the phytoextraction of metals from soil by their accumulation in theaboveground biomass involved mainly implementation of metal transporters, improvedproduction of enzymes metabolism and production of metal-detoxifyingchelators-metallothioneins and phytochelatins. Plants producing exogenous reductaseand lyase genes can transform toxic ions to harmless forms that could be volatized fromthe leaf surface or stabilized in their inner tissues, resulting in significant enhancementof plant tolerance, uptake, and detoxification performances on contaminants.In this study, the isolated Chinese cabbage anti-heavy metal genes OASA4andγ-GCS were applied to construct the plant expression vectors of pBI121-OASA4-ORF ofand pBI121-γ-GCS-ORF, respectively. Utilizing the Agrobacterium-mediated Leaf DiskMethod to transfer the plant expression vectors into tobacco variety K326, throughpre-culture, co-culture, selective culture, and subculture, transgenic tobacco buds wereabstained, and then after rooting culture, transgenic tobacco plants were gained. ThePCR results showed that, we totally got18OASA4transgenic tobacco plants and15γ-GCS transgenic tobacco plants. After4weeks of pot culture, the transgenic andnon-transgenic tobacco (as control) plants were cultured and stressed by100μmol/LCd2+. Reverse PCR of transgenic tobacco plants results showed that the target geneshave expressed in tobaccos.Assay of physiological indexes showed that the transgenic tobacco plants havehigher activities in plant resistance indicators of MDA, peroxidase, superoxidedismutase and proline, compared with non-transgenic tobacco plants; Heavy metal cadmium content determination result displayed that, cadmium adsorption capacity andcumulative capacity in transgenic tobaccos have greatly improved than thenon-transgenic tobaccos, accumulation of cadmium is mainly in the roots; tobaccocadmium accumulation is relatively small in leaves, but the transgenic tobaccos showeda certain advantages than non-transgenic tobaccos in accumulating cadmium.