| Saffron is a precious medicinal plant. Tissue and molecule levels were studied in the dissertation. Firstly, with the aim of rapid propagation of saffron, a system has been established to propagate rapidly by corm and the conditions can be controlled. Secondly, a cDNA library has been constructed from the saffron's stigma that is the medical parts. The following results were obtained.1. The new-regenerated corm was good material for the study. The whole culture was divided into three phages: initial culture, subculture and regenerate culture. The initial culture medium was MS +lmg/L 2,4-D+0.5mg/L +BA and 7% CM. The subculture medium was MS+5mg/L NAA+5mg/L BA. And the regenerate culture medium was MS + 0.5mg/L NAA+3mg/L 6-BA. Tufty buds could be induced after one or two subcultures. The frequency of induction was as high as 74%. CM may have main effect on the initiation of the explants in early period and 2,4-D and BA have synergetic effect. But it seemed that 2,4-D could not cooperate with NAA and BA with relatively high concentration. With the development of the explants, the endogeous hormones were changing accordingly. 2,4-D now may suppress the induction of tufty buds.2. The condition of illumination had a great effect on the multiplication and regeneration of tufty buds. All the tufty buds could multiply in darkness but only19% could multiply in illumination, which indicated that darkness was essential for multiplication. Tufty buds, culturing in the regenerate culture medium, could not regenerate corm in darkness but in illumination obtaining a regenerate rate of 73.3% after one-month culture. The results indicated that illumination accelerated regenerate and darkness accelerated multiplication.3. The isolation of total RNA of the stigma of saffron was a method of guanidinium isothiocyanate extraction, by which protein and pigment and so on could be got rid of efficiently and RNA did not decompose. The bands of 18S and 28S are clear after agarose gel electrophoresis. Isolated mRNA by the method of Oligo(dT) hybridization. Oligo(dT) was linked to polystyrene and could hybridize the poly(A) tail of mRNA. The mRNA washed from the spin column had no rRNA bands and didn't decompose.4. A ZAP express cDNA library was constructed by using mRNA from the stigma of saffron. By Oligo(dT)18 Linker primer containing Xho I restriction site, the mRNA of stigma was reverse transcripted into first-strand cDNA with Strata Script RT. After the second strand cDNA replacement synthesized, the uneven termini of the double- stranded cDNA were filled in with Puf DNA polymerase and EcoR I adapter were ligated to the blunt ends. Then the double- stranded cDNA was digested with Xho I restriction enzyme. Filtrating the cDNA by CL-2B gel column to get rid of small fragment. The purified cDNA was Qcloned between the EcoR I and Xho I restriction sites of the Uni-ZAP XR vector. The cDNA library was packaged in vivo into virion particles with packaging protein extracts. It was demonstrated that library contained 6.7 X105 recombinant phages and the library contained approximately 76% recombinant phages by clear/blue selection with IPTG and X-gal.
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