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使用Cav.的高效花浸转化方法

Efficient floral dip transformation method using on Cav.

作者信息

Purwantoro Aziz, Irsyadi Muhammad Burhanuddin, Sawitri Widhi Dyah, Fatumi Nor Chamidah, Fajrina Shania Nur

机构信息

Department of Agronomy, Faculty of Agriculture, Universitas Gadjah Mada, Rd. Flora, Bulaksumur, Sleman 55281, Yogyakarta, Indonesia.

Study Program of Master in Plant Breeding, Faculty of Agriculture, Universitas Gadjah Mada, Rd. Flora, Bulaksumur, Sleman 55281, Yogyakarta, Indonesia.

出版信息

Saudi J Biol Sci. 2023 Jul;30(7):103702. doi: 10.1016/j.sjbs.2023.103702. Epub 2023 Jun 1.

Abstract

Yellow cosmos ( Cav.) is a specific flowering plant and considered a suitable genetic engineering model. -mediated plant transformation is commonly used for plant genetic engineering. Floral dip transformation is one of the plant genetic transformation methods, and it involves dipping flower buds into an suspension. Studies on floral dip transformation of yellow cosmos have never been reported. Therefore, an efficient method in plant genetic engineering must be established. This study developed an effective and efficient floral dip transformation method for yellow cosmos. In this study, flower buds with sizes of 5-7 mm were used. Several parameters have been observed to optimize the floral dip method. These parameters included the optical density (OD) of culture, concentration of surfactant, and duration of flower bud dipping into the suspension. The results showed that the floral dip method was most efficient when the flower buds were dipped into suspension with OD = 0.8 and containing 5% sucrose and 0.1% Silwet L-77 for 30 s. This method enhanced the transformation efficiency at a rate of 12.78 ± 1.53%. The II and green fluorescent protein genes with sizes of 550 and 736 bp, respectively, were confirmed by polymerase chain reaction. In addition, the transgenic plants were kanamycin resistant and fluorescent under ultraviolet light observation. This finding suggests that the proposed floral dip transformation provides new insights into efficient plant genetic engineering methods for yellow cosmos.

摘要

黄秋英(Cav.)是一种特定的开花植物,被认为是一种合适的基因工程模型。农杆菌介导的植物转化常用于植物基因工程。花浸染转化是植物遗传转化方法之一,它涉及将花蕾浸入农杆菌悬浮液中。关于黄秋英花浸染转化的研究从未有过报道。因此,必须建立一种高效的植物基因工程方法。本研究开发了一种有效且高效的黄秋英花浸染转化方法。在本研究中,使用了大小为5 - 7毫米的花蕾。观察了几个参数以优化花浸染方法。这些参数包括农杆菌培养物的光密度(OD)、表面活性剂浓度以及花蕾浸入农杆菌悬浮液的持续时间。结果表明,当花蕾浸入OD = 0.8、含有5%蔗糖和0.1% Silwet L - 77的农杆菌悬浮液中30秒时,花浸染方法效率最高。该方法使转化效率提高了12.78 ± 1.53%。通过聚合酶链反应确认了分别为550和736 bp大小的GUS II和绿色荧光蛋白基因。此外,转基因植物对卡那霉素具有抗性,并且在紫外光观察下发出荧光。这一发现表明,所提出的花浸染转化为黄秋英高效植物基因工程方法提供了新的见解。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/ad05/10362457/3069d49a9485/gr1.jpg

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