Lambardi M, Lachance D, Séguin A, Charest P J
Istituto sulla Propagazione delle Specie Legnose, Consiglio Nazionale delle Ricerche, via Ponte di Formicola 76, I-50018 Scandicci (Firenze), Italy Fax: +39-55-755121 e-mail:
Natural Resources Canada, Canadian Forest Service, Laurentian Forestry Centre, 1055 du P. E. P. S., P.O. Box 3800, Sainte-Foy, Quebec G1V 4C7, Canada, , , , , , CA.
Plant Cell Rep. 1998 Dec;18(3-4):198-202. doi: 10.1007/s002990050556.
Embryonal-suspensor tissue (EST) of Mediterranean cypress (Cupressus sempervirens L.) was tested for microprojectile-DNA delivery (by the PDS-1000/He device) for different subculture periods (9, 15, and 21 days) using the plasmid vectors pRT99GUS [containing the β-glucuronidase (GUS) and neomycin phosphotransferase (NPT II) genes, and the CaMV 35S promoter], pBI426 (with a GUS::NPT II fusion gene under the control of a duplicated 35S RNA promoter), and pCGUδ0 (containing the GUS gene with the ubiquitin intron, under the control of the sunflower ubiquitin promoter). The relative strengths of the promoters as determined by GUS assays were sunflower ubiquitin>35S-35S-AMVE>35S. The highest expression level was observed when 15-day-subcultured EST was bombarded with the pCGUδ0 gene construct, which also showed high activity of the chloramphenicol acetyltransferase and NPT II genes. Green fluorescent areas were observed on EST when bombarded with the p35S-GFP plasmid, carrying the gene for the green fluorescent protein from the bioluminescent jellyfish Aequorea victoria.
利用质粒载体pRT99GUS[含有β-葡萄糖醛酸酶（GUS）和新霉素磷酸转移酶（NPT II）基因以及花椰菜花叶病毒35S启动子]、pBI426（在重复的35S RNA启动子控制下带有GUS::NPT II融合基因）和pCGUδ0（在向日葵泛素启动子控制下含有带有泛素内含子的GUS基因），对地中海柏木（Cupressus sempervirens L.）的胚柄组织（EST）进行了不同继代培养时期（9天、15天和21天）的微粒轰击DNA导入（通过PDS-1000/He装置）试验。通过GUS分析确定的启动子相对强度为向日葵泛素>35S-35S-AMVE>35S。当用pCGUδ0基因构建体轰击15天继代培养的EST时，观察到最高表达水平，其氯霉素乙酰转移酶和NPT II基因也表现出高活性。当用携带来自发光水母维多利亚多管水母绿色荧光蛋白基因的p35S-GFP质粒轰击EST时，在EST上观察到绿色荧光区域。
