| Normal growth and development of of transgenic plants are the basic premise as a kind of bioreactor, studies showed that exogenous gene had some influence on receptor plants growth. In this paper, the T0 generation pasture of Lotus corniculatus L. with Ch IFN-? gene as experimental materials to study agronomic traits, morphological,physiological and biochemical indices and the expression of genes related to growth, to explore the influence and mechanism of transfered Lotus corniculatus L.with Ch IFN-? gene in plants development, which laying a strong foundation for further exploitation and utilization of Lotus corniculatus L. Bioreactor. This study had obtained the following results:1. Identification and analysis of transgenic Ch IFN-? gene lotus plant(Lotus corniculatus L.). According to the results of histochemical staining?PCR detection?RT-PCR detection and ELISA analysis, that the foreign gene of Ch IFN-? was stably integrated in the genome of Lotus corniculatus L., and r Ch IFN-? was detected for the highest content of 0.175?g/kg in the leaves from transgenic Lotus corniculatus L.2. Studies of agronomic traits and growth of transgenic Lotus corniculatus L. Results showed that growth of transgenic lines were better than that of wild type plants(p<0.05) during the process of adventitious root formation,especially in hydroponic seedling of 14 d. The statistics of internode number showed that no significant difference were appeared between the TP and WT plants, after cultured in soil from 7d to 28 d. The biomass of the early growth stage, leaf characteristic(length, blade width, area and shape index of leaf) field growing were also no obvious difference. However, transgenic plants with more flowering time of 11 days than that of WT Lotus corniculatus L., and its first-flowering date was earlier 9 days than WT plants.3. Cytological study of transgenic Lotus corniculatus L. with Ch IFN-? gene. Based on the observation of paraffin section of Lotus corniculatus L., there were no obvious difference among the epidermis, cortex, vascular cylinder, mesophyll and veins in root, stem and leaf between the TP and WT plants. However, bits of variationswere appeared in transgenic lines, such as the collenchyma of leaf vein, phloem parenchyma cells and medullanumber, etc.4. Studies of physiological characteristics of transgenic Ch IFN-? gene Lotus corniculatus L.. Results showed that there were no obvious differences between in TP palnts and WT plants(p<0.05), including chlorophyll a content, chlorophyll b content, carotenoid content, photosynthetic pigment content, chlorophyll a/b and photosynthetic rate, etc. Foreign gene of Ch IFN-? did not affected the photosynthesis ability of receptor plant.5. Studies of low-temperature tolerance and its physiological characteristics of transgenic lotus plant.According to the results, transgenic plants with Ch IFN-? gene have higher cold resistance compare with WT lines.Further study showed that TP lines regulated these process by keeping lower MDA content(P<0.01) and higher SP,SOD, Pro contents(P<0.01).6. Studies of root system growth and its regulated mechanisms of transgenic lotus plant. Results showed that root system of TP plants had an earlier growth and better development with longer root than wild type(p<0.05),especially from 15 days of hydroponic seedling. 0.4 mg/L of IBA can effectively promote this root development in TP lines(p<0.05) by enhancing the effects of r Ch IFN-?, probably.According to the root activity analysis, TP lines had stronger root vigor than wild type, especially in 20 days of hydroponic seedling. The root primordium forming of TP was earlier 3-5 days than that of WT plants by microscopic observation. Real-time PCR analysis further suggested that the expression of LRP1 which as a marker gene for root primordium initiation was higher than in wild type(p<0.05). CKI?IAR1 and PIN1 genes which related to root growth and development from TP lines were all upward-regulated significantly than wild type(p<0.05) in the same treatment and growth stage. Ch IFN-? gene may through regulation the expression of above genes to promote the root system development in TP Lotus corniculatus L.In general, the expression of Ch IFN-? gene had no obvious effects on the stalk pitch?biomass?leaf shape and photosynthesis ability in Lotus japonicus. recipient. However, TP lines expressed better development, well-grown root system, prolonging florescence and higher cold resistance in this study. These datas will provide valuable references for Lotus corniculatus L. bioreactor expressed Ch IFN-? protein, it also provides some evidences for the plant development regulation using animal genes. |
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