信息摘要：
2026年，河南农业大学园艺学院团队在《The Plant Journal(ISSN 1365-313X)》(SCI期刊，Wiley出版社)发表文献《MhSHINE2-like interacts with MhGRF3 to promote drought tolerance via modulating stomatal aperture in apple》，文献中使用LUYOR-3415RG双波长荧光蛋白激发光源对GFP标记的转基因苹果植株进行荧光激发与成像观察，用于转基因阳性植株鉴定，为苹果抗旱基因功能研究及抗旱种质创制提供关键实验证据。
河南农业大学将LUYOR-3415RG用于转基因苹果植株GFP荧光鉴定与抗旱基因功能研究
2026年，河南农业大学园艺学院团队在《The Plant Journal(ISSN 1365-313X)》(SCI期刊，Wiley出版社)发表文献《MhSHINE2-like interacts with MhGRF3 to promote drought tolerance via modulating stomatal aperture in apple》，文献中使用LUYOR-3415RG双波长荧光蛋白激发光源用于转基因苹果植株的GFP荧光鉴定实验，对绿色荧光蛋白标记的转基因阳性植株进行荧光激发与观察，用于过表达、RNAi沉默等转基因株系的鉴定与筛选。LUYOR-3415RG双波长荧光蛋白激发光源在实验流程中承担关键的荧光激发与成像环节，通过特定波长激发光使GFP标记的转基因植株发出明亮绿色荧光，实现阳性株系的直观、快速鉴定，为研究结果的可重复性与可靠性提供保障。
河南农业大学位于河南郑州，是农业农村部、国家林业和草原局与河南省人民政府共建的省属重点高校，也是河南省特色骨干大学建设高校、河南省高校。干旱是制约全球农业生产的重要环境因子，苹果(Malus domestica Borkh)主要栽培于干旱半干旱地区，生长季干旱严重威胁苹果产业发展。该研究团队以抗旱性强的苹果野生种质湖北海棠(Malus hupehensis)为材料，通过转录组分析鉴定出干旱与ABA诱导的AP2/ERF转录因子MhSHINE2-like，利用LUYOR-3415RG的GFP荧光鉴定结合qRT-PCR分析获得过表达、RNAi等系列转基因苹果植株，并通过酵母单杂交(Y1H)、双荧光素酶(LUC)、GUS活性、凝胶迁移(EMSA)、双分子荧光互补(BiFC)、免疫共沉淀(Co-IP)等实验，揭示了MhSHINE2-like通过直接结合ABA合成关键基因MhNCED3启动子中的GAGA富集元件促进其转录，并与14-3-3蛋白MhGRF3互作、协同增强MhNCED3表达与ABA积累、调节气孔开度，从而提高苹果抗旱性的分子机制。LUYOR-3415RG的GFP荧光成像作为鉴定转基因阳性植株的关键手段，为上述研究提供了重要实验证据。
本实验采用LUYOR-3415RG双波长荧光蛋白激发光源作为转基因苹果植株GFP荧光鉴定的核心激发光源，用于对绿色荧光蛋白标记的转基因植株进行荧光激发与成像观察。载体构建与遗传转化阶段，将MhSHINE2-like、MhGRF3、MhNCED3的编码序列(CDS)分别克隆至pCAMBIA1305-GFP等载体，通过农杆菌介导法转化湖北海棠、番茄(Micro-Tom)及苹果“Orin"愈伤组织；阳性鉴定阶段，转化植株经LUYOR-3415RG激发光源辐照后观察GFP荧光信号，结合qRT-PCR分析快速鉴定阳性转基因植株，获得MhSHINE2-like过表达(OE1、OE2)、MhSHINE2-like-RNAi(RNAi1、RNAi2)、MhGRF3-OE及MhNCED3-OE等系列转基因材料；功能验证阶段，对转基因苹果植株进行断水8-12天的渐进式干旱胁迫处理，测定叶片失水率、叶绿素荧光(Fv/Fm)、电解质渗漏(EL)、丙二醛(MDA)、H2O2及O2-等生理指标，并利用Y1H、LUC、GUS、EMSA、BiFC、Co-IP、LCI等实验验证MhSHINE2-like直接结合并激活MhNCED3启动子、与MhGRF3互作并增强其蛋白稳定性，最终证实MhSHINE2-like与MhGRF3协同促进ABA积累、调节气孔开度，从而提高苹果抗旱性。LUYOR-3415RG双波长荧光蛋白激发光源用于GFP荧光鉴定的结果表明，该激发光源可稳定激发绿色荧光蛋白发光，荧光信号清晰、成像对比度高，为转基因阳性植株快速鉴定及植物抗旱基因功能研究提供了可靠的技术手段。
图1 MhSHINE2-like与MhGRF3互作验证及GFP荧光成像（论文Fig. 5）
该研究证实，LUYOR-3415RG双波长荧光蛋白激发光源可用于转基因植物GFP荧光鉴定实验，适用于荧光标记转基因植株阳性鉴定、基因功能研究与抗旱种质创制等植物学与园艺学应用场景。LUYOR-3415RG双波长荧光蛋白激发光源在国内外高校和科研院所得到广泛好评，欢迎私信或留言联系上海路阳了解详情！
原文段落
转基因苹果植株GFP荧光鉴定与遗传转化方法原文
To generate apple plants with overexpression of target genes, the pCAMBIA1305-GFP vector was digested with the restriction enzymes BamH I and Pml I (Vazyme, Nanjing, China), and the CDS of MhSHINE2-like, MhGRF3, and MhNCED3 were cloned and introduced into the pCAMBIA1305-GFP vector via ClonExpress Ultra One Step Cloning Kit V2 (Vazyme, Nanjing, China). For RNA interference (RNAi) in apple plants, a 110-bp fragment of MhSHINE2-like was introduced into the pDONOR222 vector and then transferred into the pK7GWIWG2D vector. These plasmids were separately transformed into Agrobacterium tumefaciens strain K599 for apple genetic transformation. Agrobacterium-mediated transformation of M. hupehensis was performed according to Meng et al. (2019). GFP fluorescence (LUYOR-3415RG) and qRT-PCR analysis were performed to identify the positive transgenic plants.
抗旱表型与分子机制结果原文
To investigate the biological function of MhSHINE2-like in drought stress tolerance, we first generated two MhSHINE2-like-overexpressing (OE1 and OE2) transgenic apple plants, which exhibit elevated MhSHINE2-like transcript levels (Figure S5), for subsequent drought tolerance assays. In comparison with the empty vector (EV) plants, the OE plants displayed improved tolerance to drought stress, as evidenced by the lower number of wilted leaves (Figure 1a; Figure S6) and lower water loss rate (Figure 1b). Under normal growth conditions, no significant difference was observed in chlorophyll fluorescence and Fv/Fm values between EV and OE apple leaves. However, under drought stress, the Fv/Fm values of the EV apple leaves were lower than those of OE apple leaves (Figure 1c,d). Furthermore, several related-stress indicators, including electrolyte leakage (EL), malondialdehyde (MDA), and hydrogen peroxide (H2O2), O2−, were markedly reduced in OE plants compared to EV plants under drought stress conditions (Figure 1e–h).
研究结论原文
In summary, we propose a working model that illustrates MhSHINE2-like interacts with MhGRF3 in enhancing ABA-regulated drought tolerance in apple (Figure 8). MhSHINE2-like positively regulates drought tolerance by transcriptionally activating the expression of MhNCED3. MhGRF3 enhances the protein stability of MhSHINE2-like, and the interaction between MhSHINE2-like and MhGRF3 substantially enhances the regulatory effect of MhSHINE2-like on MhNCED3, thereby promoting ABA accumulation, inducing stomatal closure, and ultimately improving drought tolerance.
图2 MhSHINE2-like过表达与RNAi苹果植株干旱胁迫表型及生理指标（论文Fig. 1）
图3 MhSHINE2-like与MhGRF3协同促进MhNCED3表达与ABA积累（论文Fig. 7）
DOI:10.1111/tpj.70761
