| Slow-growth conservation based on plant tissue culture is the most widely used method of in vitro conservation of plant germplasm resources. The target of slow-growth conservation is to restrict the growth of in vitro plantlets and reduce the frequency of subculture under the pre-request of the maintenance of genetic stability. In this research, conservative effects of different supplements to medium and different culture conditions on conservation were studied in cultivars of 'Huoju' and 'Jinba'. Moreover, genetic stability of regenerated plantlets were examined by morphology, isoenzyme and molecular markers analysis. The main results were as follows:1. Effects of abio-nutrition on in vitro conservation of chrysanthemum plantlets1/4MS or 1/2MS + 0.3mg.L-1 6-BA + 0.1mg.L-1 NAA + 30g.L-1 sucrose + 7g.L-1 agar could inhibit growing of 'Huoju' plantlets, and 100% plantlets survived after had been conserved for 12 months at (25±2)℃, 2000～3000 1x and 12h photoperiod culture conditions. But for 'Jinba', inhibit effects were't significant.2. Effects of plant growth retardant substances on in vitro conservation of chrysanthemum plantletsAt normal culture conditions, protration medium MS + 0.3mg.L-1 6-BA + 0.1mg.L-1 NAA + 30g.L-1 sucrose + 7g.L-1 agar supplemented with 1250～2000 mg.L-1 CCC or 200～600 mg.L-1 B9 or 35～140 mg.L-1 SA could prolong the conservation period of 'Huoju' and 'Jinba' plantlets to more than 10 months. Especially, optimal concentration of CCC, B9, SA is 1750, 200, 140 mg.L-1, plantlets of the two varieties had a survival rate of 74.07% and 40.74%, 81.48% and 74.07%, 100% and 100% respectively under above growth retardant substance treatments after 10-month conservation. 20mg.L-1 ABA was suitable for in vitro conservation of 'Huoju' plantlets, but for 'Jinba', the optimal concentration is 80, plantlets of the two varieties both had a survival rate of 100%, and all growed well. 3 - 5mg.L-1 PP333 or 5～9mg.L-1 PP333 could prolong conservation period for 3 months of 'Huoju' and 'Jinba' plantlets, respectively. While high concentration of 7～9mg.L-1 PP333 was harmful to 'Huoju' plantlets, and inhibit effect of 3mg.L-1 to 'Jinba' plantlets was't effective. 3. Effects of sucrose and mannitol on in vitro conservation of chrysanthemum plantletsAt normal culture conditions, 0～10g.L-1 sucrose could prolong the conservation period of 'Jinba' plantlets to 7 months, i.e., 3 months longer than control, but was't suitable for conservation of 'Huoju' plantlets. While 60～90g.L-1 sucrose could prolong the conservation period of 'Huoju' plantlets to more than 7 months, but for 'Jinba' planflets, the high concentration of sucrose accelerated the process of consenescence. 5～10g.L-1 mannitol in sucrose free medium could prolong the conservation period of 'Jinba' plantlets to 7 months. The protration medium supplemented with 10 or 30g.L-1 sucrose + 10g.L-1 mannitol, 10 or 20g.L-1 sucrose + 20g.L-1 mannitol could prolong the conservation period of 'Jinba' plantlets to more than 12 months. Especially, plantlets with the last two treatments had high survival rate of 50.00% and 60.00% after had been conserved for 12 months. Neither replacing sucrose with mannitol nor cooperating with sucrose, were suitable for in vitro conservation of 'Huoju' plantlets.4. Effects of different low temperatures on in vitro conservation of chrysanthemum plantletsStems of 'Huoju' and 'Jinba' plantlets without leaves in the protration medium conserved at 4℃directly, stems of 'Huoju' plantlets without leaves and 'Jinba' plantlets with leaves in the protration medium conserved at 10℃directly, stems of 'Huoju' plantlets no matter with leaves or without leaves in the protration medium directly conserved at 15℃and stems of 'Jinba' plantlets with leaves with 15d' pre-treatment at normal conditions before conserved at 15℃could slower the growth of plantlets, height of plantlets decreased distinctly, and had highter survival rate of 74.07% and 77.78%, 100% and 74.07%, 85.19% and 81.48% after had been conserved for 12 months.5. Effects of abio-nutrition, PP333 and CCC on in vitro conservation of chrysanthemum plantlets at 10 and 15℃Stems of 'Huoju' plantlets without leaves on protration medium conserved at 10, 15℃for 12 months had a survival rate of 100%, and growed well. Plantlets of the two cuitivars had a survival rate, similar to that control conserved on the protration medium supplemented with 3mg.L-1 PP333 at 10℃, and growed very well. Plantlets of 'Huoju' had hight survival rate, lower height, greener leaves after had been conserved for 12 months on the protration medium supplemented with 1750 mg.L-1 CCC at 10～15℃, but for 'Jinba', survival rate descended distinctly at the same conditions. 6. Genetic stability of regenerated plantletsAfter cultured in recovering medium, all plantlets conserved with the above methods recovered normal growth ability. In the regenerated plantlets, there is no obvious difference in morphological characteristics, isoenzyme zymogram of peroxidase and esterase, and amplified profiles of ISSR when compared with control. It suggested that regenerated plantlets maintained genetic stability, the slow-growth is a feasible way to conserve Chrysanthemum germplasm.
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