| Part 1 The establishment of rice gene editing technology and the transformation of aldehyde dehydrogenase in riceRice is an important grain crop,and it is very important to obtain excellent rice varieties for agricultural development.The emerging CRISPR / Cas9 gene editing technology has developed rapidly in recent years.Its construction method is simple,the targeting efficiency is high and the gene editing is targeted,which makes it has great potential in breeding work.In the search for new breakthroughs in plant breeding,the extreme enzyme genes with special functions from deep sea microbes may become a great treasure trove,and these enzymes can be introduced into plants,which may have positive effects on plant growth and stress resistance or the plant can be transformed into the "factory" which can product extreme enzymes.In this study,we established the gene editing system of rice,and used CRISPR / Cas9 technique to knockout the key gene D27 which control the tiller traits of rice.The mutant plants were successfully obtained and the insertion,substitution and deletion of the base were detected in the vicinity of the target site of the T0 gene.The results showed that the expression of D27 in the mutant plant was significantly lower than that in the wild type,and the phenotypic transformation of the T1 mutant showed obvious dwarf,multi-tiller and narrower leaves.Then,aldehyde dehydrogenase gene,which is closely related to plant resistance and has the ability of detoxifying aldehydes,is selected as the breakthrough point.The aldehyde dehydrogenase gene in the deep-sea-derived Halomonas axialensis is introduced into rice using Agrobacterium tumefaciens.The transgenic aldehyde dehydrogenase gene was successfully expressed in the rice genome and it is detected in the obtained transgenic callus.In this study,the development and utilization of deep-sea microbial resources in plants has been explored,and a transgenic functional validation evaluation system is estabilishe using CRISPR / Cas9.Part 2 Diversity analysis of DMI degrading bacteriaDue to the mass production and widespread use of plastic products,phthalate esters(PAEs)compounds pollution has been around the world.Studies have shown that PAEs have carcinogenicity and teratogenicity.They not only cause tissue damage and other hazards,but also is a class of environmental hormones,which obstruct animal reproduction and fetal development.So the pollution of PAEs is urgently need to be treated.At present,the degradation of PAEs is mainly divided into two kinds of biodegradation and abiotic degradation which includes hydrolysis and photolysis.Biodegradation is the main way of degradation of PAEs due to the slow degradation rate of hydrolysis and photolysis in natural conditions.In this study,dimethyl isophthalate(DMI),one of PAEs,was selected as the degradation substrate.Two samples,in situ and in laboratary,were collected from the deep sea at 2000 m in the South China Sea.The diversity of DMI degrading bacteria in deep seawater was analyzed in two ways which are uncultured and laboratory cultured.Cluster analysis showed that the enrichment of the samples in situ and the laboratory were quite different,suggesting that this phenomenon is due to the different oxygen content.Meanwhile,due to the cultivation methods and the limitations of culture conditions,the dominant microflora of samples obtained from uncultured and laboratory cultured methods are also quite different.In this experiment,the dominant strains are belong to eight genera,which includes Sulfurovum,Corynebacterium,Thalassospira and so on.And most of these strains were reported to be able to degrade compounds such as polycyclic aromatic hydrocarbons or petroleum.The above-mentioned strains obtained some reference value for the treatment of DMI pollution. |
