A Study On Transformation Of The Pinellia Tenata Agglutinin (pta) Gene And Waxy Antisense Gene Into Rice Mediated By Agrobacterium

Rice( Oryza sativa L) is one important foodstuff crop in China and over 50% people make it primary food.Rice comes under threat and influence of biological factors during its upgrowth; thereinto the sap-sucking insects including leafhopper, planthopper, aphides and so on that suck the phloem of rice are not only the villain of the piece but also the matter made worse by their importance as vectors of plant viruses, so those insects badly affect high yield and steady yield of rice.Using pest-resistant varieties to control pests in various crop species is very economic, effective and safe compared with using pesticides. However, conventional breeding are difficult to break through in these aspects because they get the limitation of germ plasm resources.Recent decade the development of biotechnic and gene engineering at very fast speed offers important means to create newly resistant resources.It has been shown that some plant lectins have obviously restraining or deadly effect on sap-sucking insets including leafhopper, planthoppe, aphides and so on and the function of resistant pests is very significant in cotton and rice that transferred the gene of GNA .Therefore, it has been attached more importance to study and empolder plant lectins in pest-resistant breeding in crop .Pinellia pedatisecta and Pinellia ternata are traditional Chinese medicinal plant species, belonging to family Araceae .Recent insect bioassay studies showed that lectins from P pedatisecta and P ternata had significant insecticidal activities towards cotton aphides and peach potato aphides when incorporated into artifical diets at 1.2g/L and 1.5g/L respectively. The insecticidal activities of Pinellia lectins are very similar to that of GNA.Zhang Hong-yu et al have transferred the Pinellia Tenata Agglutinin (PTA )gene into rice and the result has indicated that the transgenic homozygous lines have significant insecticidal activies.It evidently inhibits the livability and viability for rice brown planthopper.With improvement of the standard of living and advancement of the technology, people demand more and more high eating quality of rice.The endosperm starch granule that relating to proportion of amylase to amylopectin composes eating quality of rice in great degree.The highest content of amylose rice has, the badest tastes.Therefore, the contents of amylose in endosperm play an important role in eating quality and braised cooking-machining quality, which is an important breeding objective of improving rice quality.The synthesis of rice endosperm amylose is controlled by granule-bound starch synthase which is coded by waxy gene that is also master of amylose synthesis in pollen and megaspore of rice.The rice waxy gene has been separated and cloned with the development of the molecular biology. Some varieties of rice were transferredthe Pinellia Tenata Agglutinin (pta) Gene and Waxy Antisense Gene Mediated by Agrobacterium and such results below were got:1.Using immature embryos of different varieties as acceptors, some hygromicine resistant plants containing pta Gene and Waxy Antisense Gene were obtained in this study;2. PCR testing indicated that most of the resistant plants contained the selective gene and the two foreign target genes, but minority just integrated selective gene and one target gene and very few integrated selective gene and lost target gene;3. Southern blotting assay for these transgenic progenies indicated that the pta gene and waxy antisense gene have been successfully introduced and integrated into the genome of the receptors, most positive plants gained by Agrobacterium tumefaciens-mediated transformation had only one single site inserted in rice genome;4. RT -PCR assay for PTA gene showed that the PTA gene had been expressed in transcription level;5. Genetic analysis of the transgenic plant offspring showed that the separating ratio was in accordance with Mendel's single gene inheritance law and the target genes could inherit to the offspring and expressed stably;6. The amylase content in the endosperm of mature seeds from some Ti transgenic plants was measured and the results showed that the amylase content in some grains reduced .The lowest was reduced to 6.3%, which was 9.8% lower than that of the untransferred grains.