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Cucumber mosaic virus-induced gene silencing in banana

机译:黄瓜花叶病毒诱导的香蕉基因沉默

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摘要

Banana (Musa spp.) is one of the world’s most important staple and cash crops. Despite accumulating genetic and transcriptomic data, low transformation efficiency in agronomically important Musa spp. render translational researches in banana difficult by using conventional knockout approaches. To develop tools for translational research in bananas, we developed a virus induced-gene silencing (VIGS) system based on a banana-infecting cucumber mosaic virus (CMV) isolate, CMV 20. CMV 20 genomic RNA 1, 2, and 3, were separately cloned in Agrobacterium pJL89 binary vectors, and a cloning site was introduced on RNA 2 immediately after the 2a open reading frame to insert the gene targeted for silencing. An efficient Agrobacterium inoculation method was developed for banana, which enabled the CMV 20 VIGS vector infection rate to reach 95% in our experiments. CMV 20-based silencing of Musa acuminata cv. Cavendish (AAA group) glutamate 1-semialdehyde aminotransferase (MaGSA) produced a typical chlorotic phenotype and silencing of M. acuminata phytoene desaturase (MaPDS) produced a photobleachnig phenotype. We show this approach efficiently reduced GSA and PDS transcripts to 10% and 18% of the control, respectively. The high infection rate and extended silencing of this VIGS system will provide an invaluable tool to accelerate functional genomic studies in banana.
机译:香蕉(Musa spp。)是世界上最重要的主食和经济作物之一。尽管积累了遗传和转录组数据,但在农学上很重要的Musa spp中转化效率低。使用常规的基因剔除方法很难进行香蕉的翻译研究。为了开发用于香蕉的转化研究的工具,我们基于感染香蕉的黄瓜花叶病毒(CMV)分离株CMV 20开发了病毒诱导基因沉默(VIGS)系统,分别是CMV 20基因组RNA 1、2和3。分别克隆到农杆菌pJL89双元载体中,并在2a开放阅读框后立即在RNA 2上引入一个克隆位点,以插入靶向沉默的基因。针对香蕉开发了一种高效的农杆菌接种方法,该方法在我们的实验中使CMV 20 VIGS载体感染率达到95%。基于CMV 20的Musa acuminata cv沉默。卡文迪许(AAA组)的谷氨酸1-半醛氨基转移酶(MaGSA)产生了典型的褪绿表型,而尖锐分枝杆菌八氢番茄红素去饱和酶(MaPDS)的沉默则产生了光漂白表型。我们展示了这种方法有效地将GSA和PDS转录本分别降低至对照的10%和18%。 VIGS系统的高感染率和延长的沉默水平将为加速香蕉功能基因组研究提供宝贵的工具。

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