Effect of different plant growth regulators on callus and adventitious shoots induction,polysaccharides accumulation and antioxidant activity of Rhodiola dumulosa

Objective:As a medicinal plant,the resource of Rhodiola dumulosa is deficient along with the large collection.For the protection and utilization of R.dumulosa,the influence of plant growth regulators(PGRs)on callus induction and adventitious shoots differentiation,polysaccharide production and the antioxidant activity were tested.Methods:Internodes of R.dumulosa were used as explants and cultured on MS medium plus different plant growth regulators(PGRs).The anti-oxidative activities of polysaccharides were evaluated using radical scavenging assays.Results:By response surface plot,0.85 mg/L N6-benzyladenine(BA),0.34 mg/L naphthaleneacetic acid(NAA)and 0.33 mg/L 2,4-dicholorophenoxyacetic acid(2,4-D)were the optimal factors for callus induction(90.03%)from internodes explants on MS medium.The fresh weight of green callus increased 47.26 fold,when callus was inoculated on MS+thidiazuron(TDZ)0.5 mg/L+NAA 2.0 mg/L.Adventitious buds regenerated from callus on the media of MS were fortified with BA 1.0 mg/L plus NAA 0.5 mg/L,and the induction rate was 40.00%.MS plus indole-3-butyric acid(IBA)1.0 mg/L produced the highest rooting rate with 10 to 15 roots in a length of 2–3 cm per shoot.The content of total polysaccharides in callus developed on MS+TDZ 0.5 mg/L+NAA 2.0 mg/L and MS+BA 1.0 mg/L+NAA 0.5 mg/L was as high as 1.72%
2.15%.At the dose of 0.5 mg/mL polysaccharides extracted from different callus induced on MS+NAA 2.0 mg/L+TDZ 0.5 mg/L or MS+BA 1.0 mg/L+NAA 0.5 mg/L or MS+BA 0.5 mg/L+2,4-D 0.5 mg/L,the ABTS radical eliminating percentages were 82.78%,80.18%and 68.59%,respectively,much higher than that of wild plant.Conclusion:A rapid micropropagation system for R.dumulosa has been developed.The combination of TDZ and NAA or BA and NAA can increase the yield of the total polysaccharides.The polysaccharides isolated from callus and whole wild plants had stronger free radicals scavenging activities,indicating that polysaccharides from R.dumulosa are the potential pharmaceutical supplements.

Biochemical investigations during in vitro adventitious shoot regeneration in leaflet explants from nodal segments of a mature Albizia procera tree

The in vitro adventitious shoot differentiation in leaflet explants of an adult tree differed from that of leaflet explants of seedlings of Albizia procera（Roxb.）Benth. reported previously elsewhere. The leaflet explants from an adult tree passed through an initial callus phase for30 days on MS medium supplemented with 3 % sucrose,2.5 l M 2,4-D followed by a subsequent adventitious shoot differentiation phase for another 30 days on half MS medium supplemented with 0.25 l M each of BA and IBA.The regeneration rate of in vitro adventitious shoots in explants from the adult tree, i.e.1.66 shoots/callus, was lower than that from seedlings, i.e. [10 shoots/callus,which was reported elsewhere. Correspondingly, the activities of nitrate reductase and peroxidase, and endogenous phenol content remained very low during in vitro adventitious shoot differentiation in leaflet explants of an adult tree possibly due to lower availability of competent stem（juvenile） cells for the process.

Non-aerated liquid culture promotes shoot organogenesis in Eucalyptus globulus Labill

Eucalyptus is very recalcitrant to in vitro culture.In this research, an efficient shoot organogenesis system was developed using 60-day-old plants of Eucalyptus globulus grown in vitro and non-aerated liquid medium to improve shoot proliferation. Cultures were initiated with hypocotyls and leaf segments from plantlets cultivated on semisolid 1/2 MS modified medium supplemented with 4.44 μM 6-Benzyladenine(BA) and 16.1 μM 1-Naphthaleneacetic acid(NAA). Calli were transferred to shoot induction medium, with either 0.5 or 2.7 μM NAA. Shoot multiplication was carried out on 4.44 μM BA + 0.5 μM NAA medium, and semisolid and non-aerated liquid systems were compared for improving shoot proliferation.Rooting of adventitious shoots was evaluated on medium containing NAA or Indole-3-butyric acid-IBA(5 and16 μM). Callogenesis was obtained from both types of explants, although shoot formation was only obtained from leaf-derived calli. Shoot proliferation on 4.44 μM BA+0.5 μM NAA resulted in the most shoots/callus.Non-aerated liquid medium was more efficient in promoting shoot multiplication(53.5 shoots/callus) than was semisolid medium(28.5 shoots/callus). Levels of phenolic compounds were significantly reduced in the shoots cultivated in liquid medium. Efficient rooting(76%) was obtained using 16 μM IBA.

Micropropagation of yellow mangosteen:a valuable endemic tree of India

We developed a method for in vitro regenera- tion of Garcinia xanthochymus （yellow mangosteen） from matured seed segments. Multiple shoots were induced on woody plant （WP） medium supplemented with cytokinins. An average of 11 shoots per explant were regenerated from mature seed segments on WP medium containing 20 μM 6-benzylaminopurine. Histological analysis revealed that hypodermal cells of seed segments were initially involved in active division, which later developed into meriste- moids, subsequently leading to the formation of shoot buds. Shoot elongation was achieved by repeated subculturing of seed explants in shoot regeneration medium. Rooting of shoots was achieved on WP medium supplemented with indole-3-butyric acid or s-naphthalene acetic acid. Plant- lets were transplanted to pots containing soil： compost （1：1） and survival rate was 90 %.

Plant Regeneration from In Vitro Cultured Hypocotyl Explants of Euonymus japonicus Cu zhi

Adventitious shoots were successfully regenerated from hypocotyl explants of in vitro cultures of Euonymus japonicus Cu zhi. Hypocotyl slices were cultured on Murashige and Skoog （MS） and B5 basal medium supplemented with varied concentration of different plant growth-regulators, e.g., α-naphthaleneacetic acid （NAA） and indole-3-butyric acid （IBA） in combination with 6-benzylaminopurine （6-BA） and kinetin. The study showed that shoots could be directly regenerated from hypocotyl explants without the intervening callus phase; MS medium was more suitable for adventitious shoots regeneration. The ability of hypocotyls segments to produce shoots varied depending upon their position on the seedlings. The highest regeneration rate was obtained with hypocotyl segments near to the cotyledon cultured on MS basal medium supplemented with 1.5 mg L^-1 6-BA and 0.05 mg L^-1 NAA （63.64%）. The regenerated shoots were readily elongated on the same medium as used for multiplication and rooted on half-strength MS basal medium supplemented with 1.0 mg L^-1 IBA and 100 mg L^-1 activated carbon. After being transferred to greenhouse conditions, 96% of the plantlets were successfully acclimatized. This regeneration system is applied for genetic transformation now.

Optimization of in vitro Regeneration System of Benihoppe Strawberry (Fragaria × ananassa Duch.) with Orthogonal Experiment Design

[Objective] This study aimed to optimize the in vitro regeneration system of Benihoppe strawberry. [Method] Based on orthogonal experiment, the effects of different medium types, plant growth regulator types and concentrations on the regeneration of adventitious shoots of Benihoppe strawberry leaves were in- vestigated. [Result] IBA is an important factor affecting the multiplication of adventitious shoots of Benihoppe strawberry leaves; the optimal induction medium is MS ＋ 2.0 mg/L TDZ ＋ 0.2 mg/L IBA, with an induction rate of 90.00% ; the optimal differentiation medium is MS ＋ 2.0 mg/L 6-BA ＋ 0.2 mg/L IBA, with a differentiation rate of 8.1% ; the optimal multiplication medium is MS ＋ 1.0 mg/L 6-BA ＋ 0.2 mg/L IBA, with an average multiplication rate of each adventi- tious shoot of 7.67 ; the optimal subculture medium is MS ＋ 0.5 mg/L 6-BA ＋ 0. 1 mg/L 2, 4-D, with an average seeding height in each treatment of 5.98 cm; the optimal rooting medium is 1/2MS ＋ O. 5 mg/L 6-BA ＋ 0. 1 mg/L 2, 4-D, with an average number of roots in each treatment of 4.5. [ Conclusion] This stud- y laid the foundation for further investigating the genetic transformation of strawberry and improving the quality of strawberry at the genetic level.

Preliminary Studies on the Production of Artificial Seeds by Encapsutating Microtubers of Potato

The production of atificial seeds by encapsulating microtubers of potato included a series of processes. Initially, the micropropagation of adventitious buds and microtuber - inducing system should be established. Both high quality microtuber and synchronizing microtuber which have the same size were necessary for producing health potato artificial seeds. In our experiment, the optimum medium of adventitious shoot-inducing of potato Hutao, Kesi, and Favorita is DCR + 1.0 mg / kg BA+ 0.5 mg / kg IBA+ 500 mg/ kg LH; the optium medium of microtuber- inducing of Hutao and Kesi is DCR+5.0 mg/ kg BA+200 mg/ kg Lh+8% sugar, and the optium medium of microtuber -inducing of Favorita is DCR+ 5.0 mg/ kg BA+0.1 mg / kg NAA+200 mg/ kg LH+8% sugar. Microtubers of the same size were gained by selecting the synchronous microtubers. 2.0% sodium alginate was used to encapsulate the microtubers of potato to form the artificial seeds, and the concentration of Ca C12 is 4%. Phytohormone NAA promoted the germination of potato artificial seeds. These artificial seeds have more than 90% germinating frequency in plant hormone-free DCR medium.