Studies On In Vitro Culture And In Vitro Conservation In4Species Of Scented Plants Such As Ocimum Basilicum In Labiatae

Labiatae plants are of important values in economy and medcine. Ocimum basilicum, Thymusvulgaris, Melissa officinalis and Rosmarinus officinalis are good medicinal crops which areabundant in several kinds of aromatic oils. In this experiment, Ocimum basilicum, Thymusvulgaris, Melissa officinalis and Rosmarinus officinalis were used as the materials for the researchof tissue culture, in vitro conservation and other researches. Those researches included: the seedsand the stem segments with axillary buds of Ocimum basilicum and the stem segments withaxillary buds of Thymus vulgaris were used as the materials for the studies of establishingregeneration system, in vitro conservation and the changes of SOD, CAT and POD activity duringthe in vitro conservation; the Melissa officinalis plantlets were used as the materials for the studiesof in vitro conservation and the changes of SOD, CAT and POD activity during the in vitroconservation; the leaves of Rosmarinus officinalis were used as the materials for the studies ofinduction, propagation, cell suspension culture and in vitro conservation of the callus. The mainresearch results were as follows:1In vitro propagation of scented plants such as Ocimum basilicum inLabiatae1.1In vitro propagation of Ocimum basilicumThe seeds and the stem segments with axillary buds of Ocimum basilicum were used as thematerials for the researches of establishing tissue culture sterile system, proliferation, rooting,transplanting and so on. The results showed that the best method of surface sterilization for thestem segments with axillary buds of Ocimum basilicum was treated with75%alcohol (30s) and0.1%of HgCl2(5min), the best method of surface sterilization for the seeds was treated with75%alcohol (30s) and0.1%of HgCl2(8min); the MS medium containing0.1mg·L^-16-BA and0.3mg·L^-1NAA and0.3mg·L^-1KT, was the best proliferation medium, with the proliferationcoefficient of5.68; the best rooting medium was1/4MS medium with0.3mg·L^-1NAA and0.3mg·L^-1IBA, with the rooting rate of100%; the most suitable transplanting matrix was peat soil,with the survival rate of96.0%. 1.2In vitro propagation of Thymus vulgarisThe stem segments with axillary buds of Thymus vulgaris were used as the materials for thereseaches of establishing tissue culture sterile system, proliferation, rooting, transplanting and soon. The results showed that the best method of surface sterilization for the stem segments withaxillary buds was treated with75%alcohol (30s) and0.1%of HgCl₂(4min); the MS mediumcontaining1.7mg·L^-16-BA and0.1mg·L^-1NAA was the best for in vitro plantlets, with theproliferation coefficient of12.01; the best rooting medium was1/4MS medium with0.1mg·L^-1NAA and0.1mg·L^-1IBA, with the rooting rate of100%; the most suitable transplantingmatrix was peat soil, with the survival rate of92.0%.2Callus and cell suspension culture of Rosmarinus officinalisThe leaves were used as the materials for the studies of inducing and propagating callus, cellsuspension culture and in vitro conservation in Rosmarinus officinalis. The results showed that:the best induction medium was the MS medium supplemented with4.0mg·L^-16-BA and0.5mg·L^-1NAA, with the inducting rate of88.9%; the alternative culture of the MS mediumcontaining1.0mg·L^-12,4-D and the MS medium containing1.0mg·L^-12,4-D,0.3mg·L^-1KT and5mg·L^-1AgNO3was the best for callus proliferation, successive transfer culture, as well aslong-term subculture; the optimal inoculation amount was0.08g·mL^-1, the optimal subculturecycle was about7days, the optimal carbon source was sugar.3In vitro conservation of scented plantlets such as Ocimum basilicumin Labiatae3.1In vitro conservation of Ocimum basilicum plantletsThe plantlets of Ocimum basilicum were used as materials to evaluate the effects of differentconcentrations of PP₃₃₃, ABA and different factors (sugar, agarophyte, mannitol, PP₃₃₃) by usingorthogonal experimental design L16(45) on in vitro conservation. The results showed that the MSmedium containing20g·L^-1sugar,9g·L^-1agarophyte,10g·L^-1mannitol and7mg·L^-1PP₃₃₃wasthe best for the in vitro conservation, after treating4months, the survival rate was73.7%, theoptimal subculture period was about3months; also the MS medium containing9mg·L^-1PP₃₃₃andthe MS medium containing0.3mg·L^-1ABA were beneficial to in vitro conservation of Ocimumbasilicum.3.2In vitro conservation of Thymus vulgaris plantletsThe Thymus vulgaris plantlets were used as the materials for the studies of the effects of theMS medium containing different concentrations of PP₃₃₃and the MS medium containing20g·L^-1sugar,9g·L^-1agarophyte,10g·L^-1mannitol,7mg·L^-1PP₃₃₃for the in vitro conservation. Theresults showed that the MS medium containing20g·L^-1sugar,9g·L^-1agarophyte,10g·L^-1 mannitol and7mg·L^-1PP₃₃₃was the best for the in vitro conservation, after treatment for6months,the survival rate was36.7%, the optimal subculture cycle was about4months; also the MSmedium containing11mg·L^-1PP₃₃₃was beneficial to in vitro conservation.3.3In vitro conservation of Melissa officinalis plantletsThe plantlets were used as the materials to evaluate the effects of different concentrations ofABA and different factors (sugar, agarophyte, mannitol, PP₃₃₃) by using orthogonal experimentaldesign L516(4) on in vitro conservation in Melissa officinalis. The results showed that the MSmedium containing20g·L^-1sugar,9g·L^-1agarophyte,10g·L^-1mannitol and7mg·L^-1PP₃₃₃wasthe best for the in vitro conservation, after treating5months, the survival rate was44.4%, theoptimal subculture cycle was about3months; also the MS medium containing0.3mg·L^-1ABAwas beneficial to in vitro conservation.3.4In vitro conservation of callus in Rosmarinus officinalisThe callus were used as materials to evaluate the effects of different factors (sugar, mannitol,PP₃₃₃, oxyneurine) by using orthogonal experimental design L₁₆(4⁵) on in vitro conservation inRosmarinus officinalis. The results showed that the MS medium containing1.0mg·L^-12,4-D,12mg·L^-1PP₃₃₃,5g·L^-1mannitol,0.10g·L^-1oxyneurine and30g·L^-1sugar was the best for the in vitroconservation of Rosmarinus officinalis callus, which prolonged the subculture cycle to40d.4The changes of SOD, CAT and POD activity of scented plantsplantlets such as Ocimum basilicum during the in vitro conservationin LabiataeThe plantlets kept on the MS medium containing20g·L^-1sugar,9g·L^-1agarophyte,10g·L^-1mannitol and7mg·L^-1PP₃₃₃were used as the materials for the studies of the changes of SOD,CAT and POD activity during the in vitro conservation in Ocimum basilicum, Thymus vulgarisand Melissa officinalis. The results showed that the changing trends of these enzyme activities inOcimum basilicum plantlets, Thymus vulgaris plantlets and Melissa officinalis plantlets rose atfirst and then declined. Compared with the plantlets kept on the MS medium, the SOD, CAT, andPOD activity was higher and was delayed at the peak time.