PCR technique was used for amplifying THP gene in an unknown vector with primer AFP1 and AFP2.Then THP gene was ligated to pGEM T-Vector to be the plasmid pGTHP4.The plasmid pCAMBIA1301 was digested with restriction enzyme BstEⅡ and NcoⅠ,and digestion product was separated with 1% of agarose gel,then big fragment containing promoter was isolated and purified with the Agarose Gel DNA Extraction Kit.At the same way,the plasmid pGTHP4 was digested with restriction enzyme BstZⅠ and NcoⅠ,and the small fragment containing THP gene was purified from 1% agarose gel with the Agarose Gel DNA Extraction Kit.The big fragment and the small fragment were ligated at Nco Ⅰ digested cohesive-end.The ligation product was re-ligated to be cyclic plsmid by addition to a specific adapter,resulting in the pCTH823,a expression vectorof V.volvacea.
分别采用5%海藻酸钠、2%甲基纤维素、复合防腐剂 ST 和3%海藻酸钠、2%甲基纤维素、复合防腐剂 ST 制作人工种子种皮下壳和上壳,用营养小颗粒(由1.5%海藻酸钠、5.5%蔗糖、适当的无机元素和激素、保水吸附物质 C、0.1%活性炭制成)和琼脂作为胚乳材料,用制模法制成香蕉人工种子。制成的人工种子在不再供给外来营养和有菌条件下放置,便可萌发成株,成苗率达100%。
