Direct transformation of the liverwort Marchantia polymorpha L. by particle bombardment using immature thalli developing from spores
Direct transformation of the liverwort Marchantia polymorpha L. by particle bombardment using immature thalli developing from spores
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DOI：
10.1007/s00299-008-0570-5
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发表时间：
2008-09-01
影响因子：
6.2
通讯作者：
Kohchi, Takayuki
中科院分区：
文献类型：
--
作者：
Chiyoda, Shota;Ishizaki, Kimitsune;Kohchi, Takayuki
地钱，地钱L.，属于一组基地植物，是植物生物学的一个新兴模式。本研究建立了一套制备黑曲霉孢子囊的方法。在实验室条件下，通过远红光照射促进其向生殖发育的过渡，从而对多形藻进行诱变。本文报道了一种改进的M.利用从孢子发育而来的未成熟的菌体的多形物。在选择性培养基上通过用携带赋予潮霉素抗性的潮霉素磷酸转移酶基因（hpt）的质粒转化4周获得潮霉素抗性转化体。此外，还成功地将大观霉素抗性基因aadA作为M.多形。除了hpt基因之外，aadA基因的可用性应该使M. polymorpha是一种多功能的遗传操作宿主。DNA凝胶印迹分析表明，转化的菌体携带可变数量的拷贝的转基因整合到基因组中。虽然以前的系统，使用从芽生长的菌体需要一个两步选择在液体和固体培养基中8周，这里报道的系统使用从孢子发育的菌体允许在一半的时间内产生转化体的直接选择固体培养基上，促进在这个模型植物的遗传分析。
The liverwort, Marchantia polymorpha L., belongs to a group of basal land plants and is an emerging model for plant biology. We established a procedure to prepare sporangia of M. polymorpha under laboratory conditions by promoting its transition to reproductive development by far-red light irradiation. Here we report an improved direct transformation system of M. polymorpha using immature thalli developing from spores. Hygromycin-resistant transformants were obtained on selective media by transformation with a plasmid carrying the hygromycin-phosphotransferase gene (hpt) conferring hygromycin resistance in 4 weeks. The aminoglycoside-3 ''-adenyltransferase gene (aadA) conferring spectinomycin resistance was also successfully used as an additional selectable marker for nuclear transformation of M. polymorpha. The availability of the aadA gene in addition to the hpt gene should make M. polymorpha a versatile host for genetic manipulation. DNA gel-blot analyses indicated that transformed thalli carried a variable number of copies of the transgene integrated into the genome. Although the previous system using thalli grown from gemmae required a two-step selection in liquid and solid media for 8 weeks, the system reported here using thalli developing from spores allows generation of transformants in half the time by direct selection on solid media, facilitating genetic analyses in this model plant.