激发光源LUYOR-3415RG检测烟草瞬转GFP荧光
在我国农作物体系中，烟草是重要的经济作物之一。从经济贡献来看，烟草产业是部分省份的“支柱性产业"，尤其在云南、贵州、河南、湖南等主产区，烟草种植覆盖近千万农户，直接关联烟农收入、地方财政与就业稳定。烟草是典型的“喜温、喜光、喜肥"作物，对生长环境要求十分苛刻。在现代科研领域中，本氏烟草（Nicotiana benthamiana）是实验室中重要的模式植物。本氏烟草源于澳洲，容易培养和转化，目前已在各个国家广泛用于植物生物学、基因工程和分子生物学等领域的研究，例如植物与病原菌互作、代谢工程、功能基因组、合成生物学和基因编辑等。
近期，中科院华南植物园在《Plant Biotechnology Journal》期刊（IF=10.5，中科院一区top期刊）发表文献《A novel geminivirus-derived 3' flanking sequence of terminator mediates the gene expression enhancement》，文献中使用荧光蛋白激发光源LUYOR-3415RG检测烟草叶片瞬转GFP荧光，并通过专用滤镜拍摄烟草GFP荧光表达照片。LUYOR-3415RG使用便捷、检测GFP荧光速度快，大大提高了烟草荧光检测效率，为双生病毒衍生元件的功能验证提供了直观可靠的技术支撑。
荧光蛋白激发光源LUYOR-3415RG可快速检测植物基因瞬时表达与稳定表达，不用采摘叶片，可直接检测植物活体。研究团队构建SBG51系列表达载体后，通过农杆菌介导的本氏烟叶片浸润实验，利用荧光蛋白激发光源LUYOR-3415RG在2-8天的不同时间点进行荧光观测。荧光蛋白激发光源LUYOR-3415RG配备495nm专用滤光片，能高效过滤背景杂光，清晰捕捉到SBG51序列介导的GFP荧光增强效应，直观呈现出它与普通载体在荧光强度和持续时间上的差异 ——SBG51-GFP样本的荧光信号在接种后第8天仍清晰可见，而对照组在第6天已因基因沉默消失。
原文摘要
Exploring the new elements to re-design the expression cassette is crucial in synthetic biology. Viruses are one of the most important sources for exploring gene expression elements. In this study, we found that the DNA sequence of the SBG51 deltasatellite from the Sweet potato leaf curl virus (SPLCV) greatly enhanced the gene expression when flanked downstream of the terminator. The SBG51 sequence increased transient GFP gene expression in Nicotiana benthamiana leaves by up to ~6 times and ~10 times compared to the gene expression controlled by the UBQ10 promoter and 35S promoter alone, respectively. The increased GFP gene expression level contributed to the continuous accumulation of GFP protein and GFP fluorescence until 8 days post-inoculation (dpi). The SBG51 sequence also enhanced the gene expression in the transgenic Arabidopsis plants and maintained the spatio-temporal pattern of the FLOWERING LOCUS T (FT) and TOO MANY MOUTHS (TMM) promoters. We identified a123 bp of AT-rich sequence containing seven “ATAAA" or “TTAAA" elements from the SBG51DNA, which had the gene expression enhancement effect. Furthermore, the artificial synthetic sequences containing tandem repeated “ATAAA" or “TTAAA" elements were
sufficient to increase the gene expression but did not alter the polyadenylation of mRNA, similar to the function of matrix attachment regions (MAR). Additionally, the compact artificial synthetic sequence also had an effect on yeast when the expression cassette was integrated into the genome. We conclude that the geminivirus deltasatellite-derived sequence and the “ATAAA"/“TTAAA" elements are powerful tools for enhancing gene expression.
GFP fluorescence photograph and confocal microscopy . The N. benthamiana leaves infiltrated by the GFP expression vector were photographed under (hand-held) light (LUYOR-3415RG) using the 495 nm filter (LUV-495A). The GFP fluorescence of the transgenic Arabidopsis leaves, the
N. benthamiana leaves 48 h after in filtration, or the protoplasts 24 h after transfection were detected using the LEICA SP8 STED 3X fluorescence microscope confocal system.
原文图片
该研究证实，源自双生病毒δ卫星的SBG51序列可使基因表达量提升10倍。荧光蛋白激发光源LUYOR-3415RG助力中科院华南植物园基因研究，明确了 "ATAAA"/"TTAAA" 元件的增强功能，为合成生物学提供了高效便捷的检测工具。除烟草外，荧光蛋白激发光源LUYOR-3415RG还可以检测拟南芥、棉花、大豆、草莓等各类植物和作物。
荧光蛋白激发光源LUYOR-3415RG是一款便携式、充电电池供电的双波长荧光激发光源，客户可以根据需要选配两种光源，单波长6×3W LED发光，照射面积大。LUYOR-3415RG荧光蛋白激发光源可有效观察GFP、eGFP、DsRed、mCherry、tdTomato等绿色荧光蛋白和红色荧光蛋白的表达。LUYOR-3415RG在国内外高校和科研院所得到广泛好评，在全球已有近千篇引用文献。
文献DOI：10.1111/pbi.14561
