20190920「新科技」植物轉殖技術 Nature biotechnology

Chloroplast-selective gene delivery and expression in planta using chitosan-complexed single-walled carbon nanotube carriers

Seon-Yeong Kwak, Tedrick Thomas Salim Lew, Connor J. Sweeney, Volodymyr B. Koman, Min Hao Wong, Karen Bohmert-Tatarev, Kristi D. Snell, Jun Sung Seo, Nam-Hai Chua & Michael S. Strano

Abstract

Plant genetic engineering is an important tool used in current efforts in crop improvement, pharmaceutical product biosynthesis and sustainable agriculture. However, conventional genetic engineering techniques target the nuclear genome, prompting concerns about the proliferation of foreign genes to weedy relatives. Chloroplast transformation does not have this limitation, since the plastid genome is maternally inherited in most plants, motivating the need for organelle-specific and selective nanocarriers. Here, we rationally designed chitosan-complexed single-walled carbon nanotubes, utilizing the lipid exchange envelope penetration mechanism. The single-walled carbon nanotubes selectively deliver plasmid DNA to chloroplasts of different plant species without external biolistic or chemical aid. We demonstrate chloroplast-targeted transgene delivery and transient expression in mature Eruca sativa, Nasturtium officinale, Nicotiana tabacum and Spinacia oleracea plants and in isolated Arabidopsis thaliana mesophyll protoplasts. This nanoparticle-mediated chloroplast transgene delivery tool provides practical advantages over current delivery techniques as a potential transformation method for mature plants to benefit plant bioengineering and biological studies.

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利用殼聚糖聚合成的單槽奈米碳纖維管作為載體來選擇性運送並在植物葉綠體內表現外源基因

摘要

植物基因體的設計工程在近期針對作物改良、要物產聘的生合成以及永續濃豔中，是一個非常重要的工具。然而，傳統的基因轉殖工程針對細胞核內基因體修改的方式，升高了對於外來基因（因為花粉傳遞的關係）而在雜草類親緣物種中增殖的疑慮。然而，由於質體（葉綠體的前驅物質的統稱）在大多數物種都是從母本遺傳而來（並不會存在於花粉中，所以不會傳播出去），葉綠體的轉殖就沒有這樣的限制。因此推升了對於針對胞器有選擇性的奈米轉殖材料的需求。在此，我們利用脂質能和胞器的膜互換並穿透的機制，合理的設計了由殼聚糖具合成的單槽奈米碳管作為攜帶轉殖基因的載體。這個單槽奈米碳管在不需要額外作物理性或是化學藥劑的補助，即可選擇性的遞送殖體DNA到不同物種的葉綠體。我們展示了遞送並且短暫性在各種不同植物（芝麻、水芥菜、菸草、菠菜），以及在阿拉伯芥原生質體（去除細胞壁的植物細胞）的葉綠體中表達的轉殖基因。這一個由奈米粒子作為媒介的葉綠體轉殖工具和目前的轉殖技術相比，可直接轉職成株植物，因此在生物工程與生物學研究上，提供了實質的優勢。