MicroTom番茄非组培快速遗传转化方法的建立

doi:10.11983/CBB25061

植物学报

MicroTom番茄非组培快速遗传转化方法的建立

王伟坤1, 傅雯倩1, 洪秋梅1, 郑杨1, 于嘉慧1, 王艳1, 刘良淼1, 杨莉1, 2, 王长春1, 2*, 郭卫东1, 2*

1浙江师范大学生命科学学院, 金华 321004; 2金华市特色经济植物生物技术重点实验室, 金华 321004

收稿日期:2025-04-08 修回日期:2025-06-06 出版日期:2025-07-02 发布日期:2025-07-02
通讯作者: 王长春, 郭卫东
基金资助:
国家自然科学基金(No.32470174)和浙江省农业新品种选育重大科技专项(No.2021C02066-9)

A Rapid In Planta Genetic Transformation System for MicroTom Tomato

1College of Life Sciences, Zhejiang Normal University, Jinhua 321004, China; 2Key Laboratory of Biotechnology on Specialty Economic Plants of Jinhua, Jinhua 321004, China

Received:2025-04-08 Revised:2025-06-06 Online:2025-07-02 Published:2025-07-02
Contact: Changchun Wang, Weidong Guo

摘要/Abstract

摘要： MicroTom番茄(Solanum lycopersicum cv. ‘MicroTom’)是一种矮化模式植物, 因其具有较短的生命周期和清晰的遗传背景而广泛应用于植物基因功能研究。传统基于组培的MicroTom番茄遗传转化技术存在效率低、转化周期长及操作复杂等缺陷, 限制了该体系的应用。该研究以2周龄MicroTom实生苗为材料, 在切除顶端子叶与真叶后, 用根癌农杆菌(Agrobacterium tumefaciens)菌液浸润下胚轴切口并诱导再生芽, 成功构建了一套高效的MicroTom番茄非组培遗传转化体系。结果显示, 切口处再生芽阳性率为28.6%, 4–5个月即可收获T0代番茄种子, T1代植株中外源基因表达比例达73.5%。与传统组培遗传转化体系相比, 该MicroTom番茄非组培转化体系转化效率显著提高, 转化时间短且操作流程简捷, 为番茄果实性状基因功能解析提供了高效的技术平台, 同时对加快番茄分子育种进程具有重要实践意义。

关键词: MicroTom番茄, 遗传转化, 非组培, 基因功能, 分子育种

Abstract: MicroTom tomato (Solanum lycopersicum cv. ‘MicroTom’) is widely used for functional characterization due to its short life cycle and clear genetic background. However, the traditional genetic transformation system of MicroTom based on tissue culture is constrained by low efficiency, long transformation period and complex operation. Based on the somatic cell reprogramming mechanism triggered by wound signaling, this study established an efficient and rapid in planta genetic transformation system for MicroTom tomato. Wound hypocotyl were created by removing apical cotyledons and true leaves from two-week-old seedlings, followed by direct inoculation with Agrobacterium tumefaciens carrying binary vector pCY-H05251-VcDAD2-EGFP (enhanced green fluorescent protein) to induce shoot regeneration. Results showed that a 28.6% PCR-based positive efficiency of regenerated shoots in the T0 generation, with seeds derived within 4–5 months post inoculation. Antibiotic and fluorescence screening revealed approximate 73.5% lines in the T1-generation expressed the fused EGFP protein. Compared to conventional tissue culture-dependent transformation systems, this protocol enhanced transformation efficiency, shortened transformation period, and simplified sterile operational procedures. The in planta genetic transformation system provides a robust platform for functional genomics studies, and significantly lowers technical barriers in tomato genetic breeding.

Key words: MicroTom tomato, genetic transformation, in planta, gene function, molecular breeding

王伟坤, 傅雯倩, 洪秋梅, 郑杨, 于嘉慧, 王艳, 刘良淼, 杨莉, 王长春, 郭卫东. MicroTom番茄非组培快速遗传转化方法的建立. 植物学报, DOI: 10.11983/CBB25061.

Weikun Wang, Wenqian Fu, Qiumei Hong, Yang Zheng, Jiahui Yu, Yan Wang, Liangmiao Liu, Li Yang, Changchun Wang, Weidong Guo. A Rapid In Planta Genetic Transformation System for MicroTom Tomato. Chinese Bulletin of Botany, DOI: 10.11983/CBB25061.

导出引用管理器 EndNote|Ris|BibTeX

链接本文: https://www.chinbullbotany.com/CN/10.11983/CBB25061

参考文献

[1]Ahmed S, Wan Azizan WAS, Akhond MAY, Juraimi AS, Ismail SI, Ahmed R, Md Hatta MA(2023).Optimization of in vitro regeneration protocol of tomato cv.MT1 for genetic transformation..Horticulturae 9, 800., 9:800-. [2]Anjanappa RB, Gruissem W(2021).Current progress and challenges in crop genetic transformation.J Plant Physiol, 261:153411-. [3]Bailey-Serres J, Parker JE, Ainsworth EA, Oldroyd GED, Schroeder JI(2019).Genetic strategies for improving crop yields.Nature, 575:109-118. [4]Barton KA, Binns AN, Matzke AJM, Chilton MDl(1983).Regeneration of intact tobacco plants containing full length copies of genetically engineered T-DNA,and transmission of T-DNA to R1 progeny.Cell, 32:1033-1043. [5]Belanger JG, Copley TR, Hoyos-Villegas V, Charron JB, O’Donoughue L(2024).A comprehensive review of in planta stable transformation strategies.Plant Methods, 20:79-. [6]Chu P, Agapito-Tenfen SZ(2022).Unintended genomic outcomes in current and next generation GM techniques: a systematic review.Plants-basel, 11:2997-. [7]Cruz-Mendivil A, Rivera-Lopez J, German-Baez LJ, Lopez-Meyer M, Hernandez-Verdugo S, Lopez-Valenzuela JA, Reyes-Moreno C, Valdez-Ortiz A(2011).A simple and efficient protocol for plant regeneration and genetic transformation of tomato cv MicroTom for leaf explants.Hortic Sci, 46:1655-1660. [8]Dan YH, Fei ZJ, Rothan C(2007).MicroTom–a new model system for plant genomics.Genes Genom, 1:167-179. [9]Dan YH, Yan H, Munyikwa T, Dong J, Zhang YL, Armstrong CL(2006).Micro-Tom–a high-throughput model transformation system for functional genomics.Plant Cell Rep, 25:432-441. [10]Guo M, Zhang YL, Meng ZJ, Jiang J(2012).Optimization of factors affecting Agrobacterium-mediated transformation of MicroTom tomatoes.Genet Mol Res, 11:661-671. [11]胡丹玲, 孙永伟(2024).病毒介导的植物基因组编辑技术研究进展.植物学报, 59:452-462. [12]Khuong TTH, Crete P, Robaglia C, Caffarri S(2013).Optimisation of tomato Micro-Tom regeneration and selection on glufosinateBasta and dependency of gene silencing on transgene copy number.Plant Cell Rep, 32:1441-1454. [13]李君, 李岩, 刘德虎(2011).植物遗传转化的替代方法及研究进展.生物技术通报, 27:31-36. [14]Li Y, Chen Y, Zhou L, You SJ, Deng H, Chen Y, Alseekh S, Yuan Y, Fu R, Zhang ZX, Su D, Fernie AR, Bouzayen M, Ma T, Liu MC, Zhang Y(2020).Micro-Tom metabolic network: rewiring tomato metabolic regulatory network throughout the growth cycle.Mol Plant, 13:1203-1218. [15]刘小花, 张岚岚, 朱长青, 陈昆松, 徐昌杰(2008).番茄矮化微型机制及其在植物功能基因组学研究中的应用.遗传, 30:1257-1264. [16]Livak KJ, Schmittgen TD(2001).Analysis of relative gene expression data using real-time quantitative PCR and the 2-ΔΔCT method.Methods, 25:402-408. [17]Lu JH, Li SS, Deng S, Wang MG, Wu YH, Li M, Dong JS, Lu SH, Su CL, Li GF, Lang ZB, Zhu JK(2024).A method of genetic transformation and gene editing of succulents without tissue culture.Plant Biotechnol J, 22:1981-1988. [18]Lu Y, Zhu HL(2022).The regulation of nutrient and flavor metabolism in tomato fruit.Vegetable Res, 2:5-. [19]McCormick S, Niedermeyer J, Fry J, Barnason A, Horsch R, Fraley R(1986).Leaf disc transformation of cultivated tomato (L.esculentum) using Agrobacterium tumefaciens. Plant Cell Rep, 5:81-84. [20]Mei GG, Chen A, Wang YR, Li SQ, Wu MY, Hu YL, Liu X, Hou XL(2024).A simple and efficient in planta transformation method based on the active regeneration capacity of plants.Plant Commun, 5:100822-. [21]Phillips RL, Kaeppler SM, Olhoft P(1994).Genetic instability of plant tissue cultures: breakdown of normal controls.Proc Natl Acad Sci USA, 91:5222-5226. [22]Toth M, Toth ZG, Fekete S, Szabo Z, Toth Z(2020).Improved and highly efficient Agrobacterium rhizogenes-mediated genetic transformation protocol: efficient tools for functional analysis of root-specific resistance genes for Solanum lycopersicum cv. Micro-Tom..Sustainability, 14:6525-. [23]Wu XY, Zhang Y, Jiang X, Ma TS, Guo YT, Wu XX, Guo GS, Cheng XF(2024).Considerations in engineering viral vectors for genome editing in plants.Virology, 589:109922-. [24]熊发前, 刘俊仙, 刘菁, 贺梁琼, 蒋菁, 唐秀梅, 黄志鹏, 吴海宁, 钟瑞春, 韩柱强, 唐荣华(2019).花生的五种改良提取方法的比较分析及其应用.分子植物育种, 17:2207-2216. [25]Yang WT, Zhai HW, Wu FM, Deng L, Chao Y, Meng XW, Chen Q, Liu CH, Bie XM, Sun CL, Yu Y, Zhang XF, Zhang XY, Chang ZQ, Xue M, Zhao YJ, Meng XB, Li BS, Zhang XS, Zhang DJ, Zhao XY, Gao CX, Li JY, Li CY(2024).Peptide REF1 is a local wound signal promoting.Cell, 187:3024-3038. [26]Yasmeen A, Mirza B, Inayatullah S(2009).In planta transformation of tomato.Plant Mol Biol Rep, 27:20-28. [27]余柯达, 叶美娟, 陈文荣, 朱凯丽, 张常晶, 郭卫东(2016).蓝莓组织提取方法的研究.浙江师范大学学报自然科学版, 39:60-64. [28]张庆祝, 韩天富(2004).植物非组培遗传转化方法研究的进展.分子植物育种, 2:85-91. [29]郑立峰(2020).基于番茄高效再生体系的柚、基因转化体系的建立.硕士论文. 福州, 福建农林大学.,:32-33.

MicroTom番茄非组培快速遗传转化方法的建立

A Rapid In Planta Genetic Transformation System for MicroTom Tomato

PDF (PC)

可视化

摘要/Abstract

引用本文

使用本文

参考文献

相关文章 15

编辑推荐

Metrics

本文评价

[1]	李晶晶, 李艳飞, 王安琪, 王佳颖, 邓成燕, 卢敏, 马剑英, 戴思兰. 菊花品种‘万代风光’再生及遗传转化体系的建立[J]. 植物学报, 2025, 60(4): 1-0.
[2]	苏晨, 牛钰凡, 徐航, 王希岭, 于英俊, 何雨晴, 王雷. 生物钟与光温环境信号互作网络研究进展[J]. 植物学报, 2025, 60(3): 315-341.
[3]	曾文丹, 严华兵, 吴正丹, 尚小红, 曹升, 陆柳英, 肖亮, 施平丽, 程冬, 龙紫媛, 李婕宇. 发根农杆菌介导的野葛毛状根遗传转化体系[J]. 植物学报, 2025, 60(3): 425-434.
[4]	李青洋, 刘翠, 何李, 彭姗, 马嘉吟, 胡子祎, 刘宏波. 甘蓝型油菜BnaA02.CPSF6基因的克隆及功能分析(长英文摘要）[J]. 植物学报, 2025, 60(1): 62-73.
[5]	李宇琛, 赵海霞, 姜希萍, 黄馨田, 刘亚玲, 吴振映, 赵彦, 付春祥. 根癌农杆菌介导的蒙古冰草稳定遗传转化体系建立[J]. 植物学报, 2024, 59(4): 600-612.
[6]	黄慧梅, 高永康, 台玉莹, 刘超, 曲德杰, 汤锐恒, 王幼宁. 硝酸盐转运蛋白NRT2在植物中的功能及分子机制研究进展[J]. 植物学报, 2023, 58(5): 783-798.
[7]	余晓敏, 王亚琴, 刘雨菡, 易庆平, 程文翰, 朱钰, 段枫, 张莉雪, 何燕红. 根癌农杆菌介导万寿菊遗传转化体系的建立[J]. 植物学报, 2023, 58(5): 760-769.
[8]	张波, 任长忠. 燕麦基因组学与分子育种研究进展[J]. 植物学报, 2022, 57(6): 785-791.
[9]	杨澜, 刘雅, 项阳, 孙秀娟, 颜景畏, 张阿英. 谷子茎尖体外遗传转化体系的建立与优化[J]. 植物学报, 2021, 56(1): 71-79.
[10]	赖先军,张义正,古英洪,颜朗. 转昆虫抗冻蛋白基因增强甘薯抗冻能力[J]. 植物学报, 2020, 55(1): 9-20.
[11]	杨德卫,王莫,韩利波,唐定中,李生平. 水稻稻瘟病抗性基因的克隆、育种利用及稻瘟菌无毒基因研究进展[J]. 植物学报, 2019, 54(2): 265-276.
[12]	李俊华,刘世宇,李成龙,韩林林,董亚辉,张晓丽,赵喜亭,李明军. 铁棍山药微型块茎遗传转化体系的建立[J]. 植物学报, 2019, 54(1): 72-80.
[13]	吴国栋, 修宇, 王华芳. 优化子叶节转化法培育大豆MtDREB2A转基因植株[J]. 植物学报, 2018, 53(1): 59-71.
[14]	夏正俊. 大豆基因组解析与重要农艺性状基因克隆研究进展[J]. 植物学报, 2017, 52(2): 148-158.
[15]	赵喜亭, 蒋丽微, 王苗, 朱玉婷, 张文芳, 李明军. 怀黄菊间接体胚受体再生体系的建立及CmTGA1的遗传转化[J]. 植物学报, 2016, 51(4): 525-532.