| Banana (Musa spp.) was not only one of the four most popular fruits in the world and an important food crops, but also had significant economic value. It often encountered hypothermia which causes incalculable loss. To speed up the new breed, being cold resistance became an urgent problem. Fujian province had rich and excellent resource of wild banana, which can provide valuable germplasm and genetic resources for breeding bananas. Therefore it was an urgent need to carry out research on wild banana micropropagation and genetic background to exploit excellent resistance genes from wild banana for propagation of cultivated banana, to offer a new approach for cultivation, to improve banana varieties, and to lay a theoretical foundation. On the basis of previous research in a laboratory, wild bananas from Jinan district, Fuzhou were used in this study as experimental materials. Analysis of ISSR genetic diversity of Rixi wild banana was mainly conducted and the optimization of in vitro propagation condition and cloning of APX family genes, quantitative expression analysis, and transformation of tobacco etc. were also carried out. The main results are as follows:1 Genetic structure of the natural population revealed by ISSR in the wild banana from Rixi Town of Fuzhou City38 samples of the wild banana leaves from Rixi town of Fuzhou were collected as materials, The technical system of ISSR marker was used, and their genetic structures were analyzed by related software STRUCTURE and NTSYS in the study. The result showed that based on 13 primers,116 stable DNA fragments were obtained, of which polymorphism band number was 70, accounting for 60.34%, which revealed a relative high level of genetic diversity. The results of NTSYS and STRUCTURE analysis showed that 38 wild materials could be divided into 6 groups. The Q-value analysis showed there was a certain exchange of genetic information in the natural populations in Rixi Township, Fuzhou.2 The effect of light quality on micropropagation of Huanxi Wild bananaOn the basis of previous research, the effect of light quality on micropropagation of HuanXi wild banana in Fuzhou City Jinan District was studied. With laboratory trans-generational saving HuanXi wild banana somaclone as material, the effects of different light quality(red, blue, green, purple, white, yellow) on micropropagation of Huanxi wild banana plantlets were studied.The results show that:1. Under the condition of white light proliferation effect of HuanXi wild banana was best,the red was better for the preservation of wild banana plantlets, and under other light the somaclone forms can produce abnormal changes.2. By measuring chlorophyll contents of different light,the white was the highest.3. Under the condition of different light quality determination of POD activity in the order:the red>the purple>the green>the yellow>the blue>the white; CAT activity was:the white> the yellow> the purple> the blue>the green>the red.3 Cloning and bioinformatics analysis of APX gene family in Huanxi wild bananaWe cloned 7 APX genes from Huanxi wild banana; there were MuAPX1-1, MuAPX1-2, MuAPX1-3, MuAPXS-1, MuAPXT-1, MuAPX4-1, and MuAPX4-2. Their CDs varied from 1024 bp to 1466 bp, encoding the amino acid between 249 and 426. Bioinformatics prediction showed that 7 of APX gene family members are non-secreted hydrophilic proteins without signal peptide, APX1-1, APX1-2 and APX1-3 protein were stable, The other four were unstable; Subcellular localization prediction showed that APX1-1, APX1-2, APX1-3 were localized in peroxisomes,APXS-1 was localized in mitochondrial matrix space, APXT-1 was localized in the mitochondrial inner membrane and plasma membrane; APX4-1 was localized in the endoplasmic reticulum; APX4-2 was located in the plasma membrane;Huanxi wild banana APXs had KatG,PRK 15061 domain and ascorbate_peroxidase specific hits, belonging to plant_peroxidase_like super family, APX1-2 was also belonging to ApbE superfamily.APX1-2, APX1-3 and APXS-1 did not exist transmembrane,Other members had inside-out multi-transmembrane structure, Phosphorylation mainly were serine while threonine and tyrosine is less. Secondary structure by Alpha helix, Beta angle, random curl and conformation, extend the Alpha helix and random curl proportion was bigger, stretching conformation was followed, proportion of Beta angle was the least. Seven membersof APX gene family had curly spiral structure, three dimensional all were approximate spherical structure. Sequence alignment and phylogenetic analysis showed that seven members of APX family of the gene homology were lower and more distant relatives and prediction APX gene family members have different functions or a synergistic effect.4 Expressions ananlysis of APXs of Huanxi wild banana at different temperature stressExpression patterns of Huanxi wild banana plantlets at different temperature stress were ananlyzed.The results showed:Seven members of APX gene family at different temperature were expressed and expression patterns were different. Expression variations of APX1-1, APX1-3 and APX4-2 was "inverted U"; Expression patterns of APXS-1 and APXT-1, reaching the extreme, showing a "down - up - down - up" model at 13℃ APX4-1 showing the pattern of" rise after fall first "; APX1-2 was "U-type", abundantly expressed in both 28℃ and 0 ℃.5 Transformtion of MuAPX1-2 gene to tobaccoIn the experiment, the sense and antisense pCAMBIA1301-Mu-APX1-2 expression vetorjs was constructed by double enzyme digestion, and successfully transformed into Agrobacterium EHA105. Genetic transformation of tabacco leaves was conducted successfully and PCR and GUS detection of resistant plants after hygromycin selection were anlalyzed to detect the transformation result in target gene.The results showed that 17 sense transformation plants and 20 antisense transformation plants were obtained.The transformation frequencies amounted to 69.81%. We also collected the sense and antisense transgenic tobacco plants seeds. |
PDF Full Text Request