北京林業(yè)大學(xué)在《International Journal of Biological Macromolecules》發(fā)表了《Genome-wide identification and functional analysis of silicon transporter family genes in moso bamboo (Phyllostachys edulis)》，文獻(xiàn)中使用了LUYOR-3415RG便攜式熒光蛋白激發(fā)光源用于觀察EGFP在擬南芥上的表達(dá)。
Silicon (Si) has crucial effects on plant development and stress resistance. Silicon transporters regulate Si absorption, transport, and distribution in plants. In this study, we identified and characterized the Si transporter gene family of moso bamboo (Phyllostachys edulis) and cloned seven putative Si transporter genes. Moso bamboo Si transporters contain conserved functional domains that mediate the accumulation of considerable amounts of Si. The analysis of gene duplication patterns and divergence times suggested that the expansion of the moso bamboo Si transporter family was mainly due to segmental duplications. The expression of moso bamboo Si transporter genes, which varied among organs, was significantly modulated by Si treatments. The subcellular localization analysis showed that Si transporters are plasma membrane proteins. The Si content increased in transgenic Arabidopsis overexpressing PeLsi1-1 or PeLsi1-2, which affected vegetative and reproductive growth. Our single-particle tracking analysis revealed the four diffusion modes of PeLsi1-1 on the plasma membrane. Moreover, the particle velocity, dwell time, and motion range of PeLsi1-1 decreased in response to Si treatments. The results of this study will further clarify the molecular mechanisms underlying Si absorption and accumulation in bamboo plants.
Genetic transformation and identification of transgenic Arabidopsis
Recombinant plasmids containing target genes were separately transferred into Agrobacterium tumefaciens strain GV3101 cells (Zomanbio, Beijing, China) via a freeze–thaw method for the subsequent transformation of Columbia-0 wild-type Arabidopsis with pBI121-35S::EGFP, PeLsi1-1, PeLsi1-2, and PeLsi2 according to the floral dip method [48]. Harvested T1 generation seeds were screened on half-strength Murashige & Skoog (MS) medium (Table S4) containing 50?mg/L kanamycin. The LUYOR-3415RG dual-wavelength fluorescent protein excitation light source (LUYOR, Shanghai, China) was used to screen for transgenic Arabidopsis lines with high EGFP expression levels (excitation and emission wavelengths were 440–460 and 500?nm, respectively). The kanamycin-resistant T1 seedlings were analyzed using the TransDirect® Plant Tissue PCR Kit (TransGen) and the EGFP-F/EGFP-R primers (Table S6). Twelve positive seedlings per transgenic Arabidopsis line were selected for the subsequent culture and examination. The T1 plants were irradiated using LUYOR-3415RG, after which three plants with high EGFP expression levels that also conformed to Mendelian inheritance were selected for each transgenic Arabidopsis line. AtActin2 (AT3G18780) was used as an internal control to identify T3 transgenic Arabidopsis plants via an RT-PCR analysis involving Si transporter gene-specific primers (Table S6).
上海路陽(yáng)有限公司生產(chǎn)銷售各種款式熒光蛋白觀測(cè)燈，有LUYOR-3280手電筒式熒光蛋白觀察手電筒、LUYOR-3415手持式雙波長(zhǎng)熒光蛋白激發(fā)光源、LUYOR-3421臺(tái)式熒光蛋白激發(fā)光源，能夠滿足觀察熒光蛋白在模式動(dòng)物、植物葉片、種子、愈傷等上的表達(dá)。
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