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转水稻几丁质酶基因香蕉(“大麦克”,AAA 基因组群)表达赋予对黑叶斑病的抗性。

Expression of a rice chitinase gene in transgenic banana ('Gros Michel', AAA genome group) confers resistance to black leaf streak disease.

机构信息

Laboratory of Tropical Crop Improvement, Department of Biosystems, Faculty of Bioscience Engineering, KU Leuven, Kasteelpark Arenberg 13, bus 2455, 3001, Leuven, Belgium.

出版信息

Transgenic Res. 2013 Feb;22(1):117-30. doi: 10.1007/s11248-012-9631-1. Epub 2012 Jul 13.

Abstract

Transgenic banana (Musa acuminata 'Gros Michel') integrating either of two rice chitinase genes was generated and its resistance to Black Leaf Streak disease caused by the fungus Mycosphaerella fijiensis was tested using a leaf disk bioassay. PCR screening indicated the presence of the hpt selectable marker gene in more than 90 % of the lines tested, whereas more than three quarters of the lines contained the linked rice chitinase gene resulting in a co-transformation frequency of at least 71.4 %. Further, a unique stable integration of the transgenes in each line revealed some false negative PCR results and the expected co-transformation frequency of 100 %. The transgene insert number per line ranged from 1 to 5 and single transgene insert lines (25 % of all) were identified. Considerable delay in disease development (up to 63 days post-incoculation) over a monitoring period of 108 days occurred in nine lines with extracellularly targeted chitinase out of 17 transgenic lines tested and their necrotic leaf area decreased by 73-94 % compared to the untransformed susceptible control line. Finally, correlation between symptom development and rice chitinase expression was confirmed in two lines by Western analysis. The potential of rice chitinase genes to enhance resistance against M. fijiensis in banana was demonstrated as well as the usefulness of the leaf disk bioassay for early disease screening in transgenic banana lines.

摘要

转基因为‘大麦克’香蕉(Musa acuminata 'Gros Michel')的植株整合了两种水稻几丁质酶基因,利用叶盘生物测定法对其抗由真菌木薯球腔菌(Mycosphaerella fijiensis)引起的黑叶斑病的能力进行了测试。PCR 筛选表明,在所测试的 90%以上的品系中存在 hpt 选择标记基因,而超过四分之三的品系含有连接的水稻几丁质酶基因,从而导致共转化频率至少为 71.4%。此外,每条品系中转基因的独特稳定整合导致一些假阴性 PCR 结果,预期的共转化频率为 100%。每条品系的转基因插入数从 1 到 5 不等,鉴定出了单转基因插入系(占所有系的 25%)。在所测试的 17 个转基因系中,有 17 个系中的 9 个系(占 53%)表现出胞外靶向几丁质酶的明显发病延迟(接种后长达 63 天),其坏死叶片面积比未转化的易感对照系减少了 73-94%。最后,通过 Western 分析在两条系中证实了症状发展与水稻几丁质酶表达之间的相关性。证明了水稻几丁质酶基因在增强香蕉对木薯球腔菌的抗性方面的潜力,以及叶盘生物测定法在转基因香蕉系早期疾病筛选中的有用性。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/781d/3525978/47315d2113d7/11248_2012_9631_Fig1_HTML.jpg

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