Growth and Transcriptomics Analysis ofMichelia macclurei Dandy Plantlets with Different LED Quality Treatments

Michelia macclurei Dandy is a significant tree species that has extensive cultivation for forestry and horticulture purposes in southern China,owing to its economic and practical importance.Light quality influences plantlet growth and development during tissue culture.However,the growth characteristic and molecular regulation of M.macclurei under different light quality conditions are not well understood yet.In this study,we investigated the morphological,chlorophyll content,and transcriptomic responses of M.macclurei plantlets under different light-emitting diode(LED)qualities,including white,blue,and red light.The results showed that blue light significantly increased plant height(21.29%)and leaf number(18.65%),while red light decreased plant height and leaf number by 7.53%and 16.49%,respectively.In addition,the plantlets’chlorophyll content and etiolation rate were significantly reduced by blue and red light quality compared to white light.Compared to white light,blue light had a negative effect,leading to decreased rooting rate(64.28%),root number(72.72%),and root length(75.86%).Conversely,red light had a positive effect,resulting in increased rooting rate(24.99%),root number(109.58%),and root length(72.72%).Transcriptome analysis identified 54 differentially expressed genes(DEGs)in three groups that consisted of blue light vs.white light(BL-vs-WL),red light vs.white light(RL-vs-WL),and red light vs.blue light(RL-vs-BL).Specifically,21,7,and 41 DEGs were identified in the three groups,respectively.The DEGs found in the RL-vs-WL and BL-vs-WL groups were involved in plant hormone signaling,nitrogen metabolism,and phenylpropanoid biosynthesis pathways,which suggests that M.macclurei plantlets adapt to the changes of light quality via modulating gene expression.Overall,our study provides valuable insights for understanding the molecular and morphological responses of M.macclurei plantlets under different light qualities.

Evaluation of Different Substrates Compositions for Acclimatization of Tissue Culture Taro Plantlets in a Propagator

Taro is cultivated in most Regions of Cameroon and it is affected by taro leaf blight disease since 2010 which has decreased its production. Lack of disease-free planting materials has been a main problem to farmers. This study was carried out at International Institute of Tropical Agriculture (IITA) Yaounde and Institute of Agricultural Research for Development (IRAD) Bambui to assess different substrates for acclimatization of tissue culture taro plantlets in apropagator. No information is available on acclimatization of Cameroonian taro plantlets in different substrates. Taro plantlets from tissue culture were acclimatised in a propagator for six weeks under different substrates, the first substrate consisted of sterile three parts of soil and one part of river sand mixed together (3:1), the second substrate consisted of sterile two parts of soil and two parts of river sand mixed together (2:2), the third substrate consisted of sterile two parts of soil, one part of rice husk and one part of river sand mixed together (2:1:1) and the fourth substrate consisted of sterile one part of soil and three parts of river sand mixed together (1:3). After acclimatisation of the different taroplantlets (Dark green petiole with small leaves (L1), Red petiole with small leaves (L2), Light green petiole with large leaves (L3) and Light green petiole with small leaves (L4) in these four substrates, it was observed that the best growth rate of plant was recorded on substrate sand + soil (1:3). The other substrates showed moderate growth of plants. Substrate sand + soil (1:3) can be recommended for acclimatization of Cameroonian taro plantlets.

Effects of Sodium Chloride Stress on Growth of Sweet Potato Plantlets in vitro and Ion Content

[Objective] This study aimed to investigate the salt-tolerance mechanism of sweet potato.[Method]Two sweet potato varieties of Xu 25-2(salt-tolerant cultivar)and Triumph 100(salt-sensitive cultivar)were treated by sodium chloride with the concentration of 0 mmol/L and 100 mmol/L.After 20 days,Na+ content and Na+/K+ ratio in the roots,shoots and leave were determined by the flame photometer,while dry weight and fresh weight of roots,shoots and leave in different varieties were also studied.[Result]The growth of two sweet potato varieties was inhibited under salt stress,so the plant became shorter,leaf and root became fewer,dry weight of roots and leave decreased,but seedlings of Xu 25-2 were inhibited slightly.Furthermore,Na+ content and Na+/K+ ratio in roots,shoots and leaves of two sweet potato varieties increased.Na+ content of salt-tolerant Xu 25-2 was low in roots,shoots and leaves,while Na+ content of salt-sensitive Triumph 100 was high in shoots and leave of seedlings,but the change range of Xu 25-2 was less than that of Triumph 100.[Conclusion]The lower Na+ content and Na+/K+ ratio in leaves under salt stress were the most important characteristics for salt-tolerance of sweet potato varieties.

Callus induction from leaves of different paulownia species and its plantlet regeneration

The experiment was carried out on five different species of Paulownia for callus induction from leaves. MS medium was adopted as basic medium, and from different combinations of NAA and BA the suitable media were determined for callus induction, bud differentiation, and root differentiation of five different species. MS+0.5NAA+4BA, MS+0.3NAA+2BA, MS+0.5NAA+4BA, MS+0.3NAA+6BA, and MS+0.3NAA+8BA were suitable media of callus inductions of leaves, respectively, for Paulownia tomentosa, Paulownia australis, Paulownia fortunei, Paulownia elongata and P. tmentosa x P. fortunei, and MS+0.3NAA+12BA, MS+0.3NAA+12BA, MS+0.5NAA+12BA, MS+0.5NAA+12BA, and MS+0.7NAA+12BA were suitable media for bud differentiation from leaf callus respectively for above five species. The rooting media was determined as 2MS+0.1NAA, 1/2MS+0.1NAA, 1/2MS, 1/2MS+0.3NAA, and 1/2MS+0.5NAA. These results provide reference data for breeding new fine va-rieties with different kinds of Paulownia protoplasts fusions.

Research on Plantlet Rooting of Lagerstroemia indica

Tissue culture plantlets of Lagerstroemia indica were used to study the effects of different auxins on in-vitro rooting, and the effects of various media in transplanting of rooted plantlets and ex-vitro rooting. For in-vitro rooting of test-tube plantiets of Lagerstroemia indica, 0.5 mg/L IBA is a suitable growth regulator; in the case of transplanting of rooted plantlets, the medium ratio of peat soil to perlite at 7：3 is better; and as to ex-vitro rooting of Lagerstroemia indica plantlets, the medium ratio of peat soil to perlite at 3：7 is more suitable.

Effect of Phytohormones on Adventitious Bud Differentiation from Bulb Scales of Oriental Lily Test-tube Plantlets

[Objective] The purpose of this study is to investigate the effects of differ- ent phytohormones on the adventitious bud differentiation of oriental lily. [Method] The bulb scales of the test-tube plantlets of Tiber, Rodina and Constanta were cul- tured in media supplemented with different cytokinin and auxin at different concen- tration, and then the adventitious buds in each treatment were calculated. [Result] Cytokinins had different influence on the adventitious bud differentiation of the three oriental lily cultivars. Among them, 6-BA had the best effect to induce the adventi- tious bud differentiation from bulb scales of Tiber and Rodina, but there was some difference in the optimal concentration. KT had the best effect to induce the adven- titious bud differentiation of Constanta. The auxins had little influence on the quality of the adventitious bud of the three oriental lily cultivars, but caused some difference in differentiation coefficients. [Conclusion] The most suitable media for the adventi- tious bud differentiation from bulb scales in vitro of Tiber, Rodina and Constanta were MS＋0.2 mg/L 6-BA＋0.2 mg/L 2,4-D, MS＋I.0 mg/L 6-BA＋0.2 mg/L IAA, MS＋ 1.0 mg/L KT＋0.5 mg/L 2,4-D, respectively.

Technology Research of Plantlets Rooting and Transplanting on Tissue Culture of Ilex centrochinensis

Plantlets of Ilex centrochinensis were used in its tissue culture to study the effect of auxin and gibberellin in vitro rooting, and the effect of various matrix in transplanting rooted plantlets and in rooting outside the tube. The highest fre- quency of rooting was obtained when growth regulator combination was IBA 0.2 mg/L ＋ GA3 0.4 mg/L in vitro rooting, and when the ratio of sand and humus soil was 5 to 5 in rooting outside the tube. When the ratio of perlite and peat soil was 3 to 7, the highest survival of rooted plantlets in transplanting was obtained. This study may provide the basis for improving the factory system of tissue culture of llex centrochinensis.

Study on the Flower Development of Testtube Plantletsof Orychophragmus violaceus

In this study, testtube plantlets of O.Violaceus were cultured on MS media and treated with low temperature, phytohormones, etc. The following results were obtained: 1. The cold treatment of 5～7℃ for more than 7 days was needed for flower initiation of the testtube plantlets originated from germinated seeds. 2. When cultured on the MS media supplemented with 2mg/L gibberellin (GA3), the testtube plantlets could initiate flowers without cold treatment. 3. When the MS media was supplemented with a reduced amount of NH4NO3, flower initiation of the testtube plantlets could be accelerated. The results of the study on flower development of O.Violaceus may be useful for the production of testtube flowers of this ornamental plant species.

Effects of Lanthanum and Europium on Rooting of Plantlet Eriobotrya Japonica Lindl. in vitro

The effect of La 3+ and Eu 3+ on the rooting of Eriobotrya japonica Lindl. plantlet in vitro was studied with adding La 3+ and Eu 3+ to the rooted medium. The rooting rate, the number of root and the length of root were studied after transplanting 20 d. The activity of peroxidase, nitrate reductase and fresh weight of roots were determined after transplanting 44 d. The results show that the optimum concentration range of La 3+ (1.0～3.0 μmol·L -1), Eu 3+ (2.0～3.0 μmol·L -1) in the rooted medium can increase the rooting rate and the fresh weight of roots, and promote the length of root and raise the activities of peroxidase and nitrate reductase significantly. La 3+ has more effect in improving the rooting rate, root length and the activities of peroxidase and less effect in promoting root fresh weight and the activities of nitrate reductase than Eu 3+.

In vitro callus induction and plantlet regeneration of Achyranthes aspera L.,a high value medicinal plant

Objective:To study callus induction from different explants(internode,leaf,root)and in vitro plantlets propagation from medicinally important plant Achyranthes aspera L.Methods:Sterilized explants were prepared by uning 0.1%HgCl_2 and 0.5%Bavistin and callus was obtained when cultured onto Murashige Skoog's(MS)medium by using different concentrations and combination of 2,4-D.NAA.BAP,IAA,IBA with 3%sucrose and 0.8%agar.Induced callus was immediately transferred to MS medium containing at different concentrations of phytohormones for shootlets and rootlets induction respectively.Results:Sterilization treatment of 0.1%HgCl_2.for 2-3 min and Bavistin 0.5%for 10-12 min showed the highest percentage of asepsis and survival rate.Maximum induction of callus was obtained from a combination of 2.0 mg/L 2,4-D and 0.5 mg/L NAA from leaf.Highest shootlets number(4.83±0.l7)and length(3.8±0.16)cm were observed on full strength MS medium when fortified with BAP 4.0 mg/L and KIN 0.5 mg/L.Concerted efforts of BAP 10 mg/L and NAA 0.5 mg/L on full strength MS medium showed highest leaf number(6.77±0.94).In vitro raised shoots were allowed to root on different strengths of MS medium fortified with IAA and IBA at different concentrations.Experimentally,3.0 mg/L IBA was enabled to induce maximum rootlets number(10.0±9.82)on full strength MS medium.Afterwards,regenerated shoots with well developed roots were successfully subjected to hardening process and were acclimatized.The survived plantlets showed 66.67%survival frequency without any morphological abnormality.Conclusions:The results demonstrated that different explants were good source of callus induction,morphology analysis as well as indirect plantlets regeneration.

Effects of different LEDs light spectrum on the growth,leaf anatomy,and chloroplast ultrastructure of potato plantlets in vitro and minituber production after transplanting in the greenhouse

Light spectrum plays an important role in regulating the growth and development of in vitro cultured potato(Solanum tuberosum L.) plantlets. The status of potato plantlets at the end of in vitro stage influences the minituber production after transplanting. With 100 μmol m^-2s^-1 total photosynthetic photon flux density(PPFD), a light spectrum study of 100% red light emitting diodes(LEDs) light spectrum(RR), 100% blue LEDs light spectrum(BB), 65% red+35% blue LEDs light spectrum(RB), and 45% red+35% blue+20% green LEDs light spectrum(RBG) providing illumination at the in vitro cultured stage of potato plantlets for 4 weeks using fluorescent lamp as control(CK) was performed to investigate the effects of LEDs light spectrum on the growth, leaf anatomy, and chloroplast ultrastructure of potato plantlets in vitro as well as the minituber yield after 2 months transplanting in the greenhouse. Compared to CK, RB and RBG promoted the growth of potato plantlets in vitro with increased stem diameter, plantlet fresh weight, plantlet dry weight, and health index. Furthermore, BB induced the greatest stem diameter as well as the highest health index in potato plantlets in vitro. Root activity, soluble protein, and free amino acid were also significantly enhanced by BB, whereas carbohydrates were improved by RR. In addition, thickness of leaf, palisade parenchyma and spongy parenchyma was significantly increased by BB and RBG. Chloroplasts under BB and RBG showed well-developed grana thylakoid and stroma thylakoid. Unexpectedly, distinct upper epidermis with greatest thickness was induced and palisade parenchyma and spongy parenchyma were arranged neatly in RR. After transplanting in the greenhouse for 2 months, potato plantlets in vitro from BB, RB, and RBG produced high percentage of large size tuber. BB improved fresh and dry weights of the biggest tuber but decreased tuber number per plantlet. In addition, RBG increased tuber number as well as tuber fresh and dry weight slightly. Our results suggested monochromatic blue LEDs as well as combined red, blue or/and green LEDs light spectrum were superior to fluorescent lamp spectrum in micro-propagation of potato plantlets. Therefore, the application of RBG was suitable;BB and RB could be used as alternatives.

Effect of mini-cutting size on adventitious rooting and morphophysiological quality of Ilex paraguariensis plantlets

As the energizing and therapeutic effects of infusions of mate(Ilex paraguariensis A.St.-Hil)leaves are increasingly recognized,rising global consumption is also increasing the demand for plantlets with high genetic and morphological qualities.Mini-cuttings are an alternative for clonal plantlets;however,more details are needed on factors that affect the success of this technique.Here,toward maximizing production,we evaluated adventitious rooting competence of mini-cuttings of different sizes and the morphophysiological quality of the resultant plantlets.Small(one bud;length up to 2.5 cm),medium(multiple buds;length 2.6–5.0 cm),and large(multiple buds;length 5.1–10.0 cm)mini-cuttings were planted in a mixture of commercial substrate,vermiculite,and coarse sand(1:1:1 v:v:v),grown in a humidity chamber,then evaluated after 45,60,75,and 90 days for survival,percentage of callus and shoot formation,number and length of shoots,percentage of rooted cuttings,and number and length of roots.Any rooted mini-cuttings were then cultivated in commercial substrate and subsurface soil(2:1 v:v)and evaluated after 30,60,90 and 120 days of cultivation for morphophysiology of the plantlets,percentage of survival,number of leaves,shoot height,stem diameter,and ratio of shoot height to stem diameter.After 120 days,the plantlets were evaluated for the ratio of shoot and root dry mass,total length,surface area,total volume,and mean diameter of roots and Dickson quality index.Adventitious rooting of the mini-cuttings and the morphophysiological quality of the plantlets were not influenced by the size of the vegetative propagule.Therefore,single-bud mini-cuttings have similar adventitious rooting competence,and result in plantlets with adequate morphophysiological quality,based upon leaf development,shoot height,stem diameter and length,and surface area,volume and diameter of roots.Single-bud mini-cuttings up to 2.5 cm long can thus be used to maximize production of mate plantlets by mini-cuttings.

Transcriptome analysis reveals effects of red and blue lightemitting diodes(LEDs)on the growth,chlorophyll fluorescence and endogenous plant hormones of potato(Solanum tuberosum L.)plantlets cultured in vitro

Red and blue light illumination has been reported to significantly affect plantlet growth.Potato is an important food and feed crop in the world and potato plantlet cultured in vitro plays an important role in potato production.However,few studies have documented the effects of red and blue light on the growth of potato plantlets revealed at the transcriptome level.The objective of this study was to determine the growth and physiological responses of potato plantlets cultured in vitro under monochromatic red(RR),monochromatic blue(BB)as well as combined red and blue(RB)LEDs using the RNA-Seq technique.In total,3150 and 814 differentially expressed genes(DEGs)were detected in potato plantlets under RR and BB,respectively,compared to RB(used as control).Compared to the control,the DEGs enriched in"photosynthesis"and"photosynthesis-antenna proteins"metabolic pathways were up-regulated and down-regulated by BB and RR,respectively,which might be responsible for the increases and decreases of maximum quantum yield(F_(v)/F_(m)),photochemical quantum yield(φ_(PSII)),photochemical quenching(q_(P))and electron transfer rate(ETR)in BB and RR,respectively.Potato plantlets exhibited dwarfed stems and extended leaves under BB,whereas elongated stems and small leaves were induced under RR.These dramatically altered plantlet phenotypes were associated with variable levels of endogenous plant hormones gibberellin(GAs),indoleacetic acid(IAA)and cytokinins(CKs),as assessed in stems and leaves of potato plantlets.In addition,monochromatic red and blue LEDs trigged the opposite expression profiles of DEGs identified in the"plant hormone signal transduction"metabolic pathway,which were closely related to the endogenous plant hormone levels in potato plantlets.Our results provide insights into the responses of potato plantlets cultured in vitro to red and blue LEDs at the transcriptomic level and may contribute to improvements in the micro-propagation of potato plantlets cultured in vitro from the light spectrum aspect.

Synthetic Seed Preparation, Germination and Plantlet Regeneration of Litchi (Litchi chinensis Sonn.)

Litchi chinensis sonn.) ranks second after mango amongst the most important fruit crops cultivated worldwide. Litchi is a very valuable crop throughout the world because it is a table fruit and wines are also produced from it. The existing cultivars are highly polyploidy and heterozygous in nature. It is propagated through air layering and marcottage methods and storability is very low. Synthetic seeds can be stored for a long time and its genetic constitution could remain the same. For germplasm maintenance and clonal propagation, synthetic seeds can be used. Somatic embryogenesis has been reported from anther or embryogenic suspension culture in various species of litchi. Regeneration via organogenesis and somatic embryogenesis from zygotic embryos has also been reported in certain species. Developing a methodology for getting somatic embryogenesis with a high frequency from zygotic embryos which is available once in a year, would be particularly useful for genetic improvement of litchi. Cotyledonary stage somatic embryos developed from zygotic embryos were encapsulated in 2% alginate gel. The encapsulated somatic embryos (ESEs) germinated successfully on 0.7% agar medium containing 3% sucrose concentration in NN basal medium (half strength of major and minor salts) with 1 mg·l-1 of gibbrellic acid. Percentage germination and plantlet development for ESEs was higher than that of non encapsulated embryos (NSEs). In comparison to different hormones, gibberellic acid has a significant influence on the germination rate of ESEs after one week of dehydration was seen maximum at 9% sucrose and abscisic acid (1 mg·l-1) in half strength of major and minor salts in Nitsch and Nitsch medium resulting in extended storage up to 90 days without loss in germination potential and capability to regenerate into plantlets. Normally developed plantlets regenerated from ESEs were successfully adapted to soil to obtain a full grown plant.

Sterile Germination of Dendrobium chrysotoxum Seeds and Rapid Propagation of Its Plantlets

[Objective] This study was to produce plant seeds on a large scale via sterile germination of the capsules of Dendrobium chrysotoxum and rapid propagation technique.[Method] A large amount of protocorm-like bodies produced from the sterile germination of D.chrysotoxum capsules,were transferred to four different kinds of bud induction media to obtain the optimal media suitable for plantlet differentiation and growth;and then with N6 as basic medium,1.0,1.5 and 2.0 mg/L of NAA and IAA were tested to obtain the optimal media suitable for rooting.[Results] On the medium of MS appended with 1 mg/L 6-BA ＋10% banana puree ＋ 20 g/L sucrose ＋6 g/L agar＋1 g/L AC,seed germination rate was up to 90%.The optimal medium for differentiation of D.chrysotoxum protocorm-like bodies was N6＋ 2 mg/L NAA ＋ 0.5 mg/L 6-BA ＋10% banana puree ＋ 20 g/L sucrose ＋ 0.5 g/L peptone ＋ 5.8 g/L agar ＋0.5 g/L AC,grown from which the plantlets were even and orderly in height;and the optimal medium for rooting was N6＋1.5 mg/L NAA ＋10% banana puree ＋ 20 g/L sucrose ＋ 5.8 g/L agar ＋1 g/L AC,grown from which the plantlets developed more,robust and orderly roots,and their leaves were in dark green color.[Conclusion] Our results provided reference for the rapid propagation of D.chrysotoxum.

Saline Stress Response of Plantlets of Common Wheat （Triticum aestivum） and Its Wild Congeners

The aim of this work was to study the reactions of wheat （T. aestivum L.） and its wild congeners under salt stress. Physiological mechanisms that underlie traits for salt tolerance plantlets have been used to test some wheat species and to identify sources of salty tolerance. Stress conditions were founded by means of exposure of the plantlets in NaCI water solution, 1.68% that demonstrably differentiates the samples by growth parameters. Under laboratory conditions the ability of seedlings to bear a significant shortage of water, their ability to accumulate biomass, changing the parameters of linear growth processes and changes in water content in leaves and roots of seedlings under stress was determined. Interest is also the rate of growth of root fibrils in saline conditions. These experiments allowed to provide a comprehensive diagnosis of studied forms of wheat and make an initial opinion on their salt tolerance. Different features of abiotic stress responses of some wheat species at the early stages of development were shown that testifies necessi~ of great number of approaches at selection of genetic material for improvement of modern wheat eultivars. This work is the first phase of research for practical breeding.

Effects of Neodymium on Nitrogen Metabolism of Plantlet Loquat in Vitro

The effect of NdCl_3 on the rooting and nitrogen metabolism of loquat in vitro was studied when NdCl_3 was added to the rooted medium. The results show that 0.4 μmol·L^(-1) NdCl_3 in the rooted medium can obviously increase the rooting rate, length of root and fresh weight of roots, and enhance the activities of nitrate reductase, glutamine synthetase, glutamate dehydrogenase in the root system and in the leaves. The transformation of NO_3^- to NH_4^+ in root system and leaves are promoted and the nitrogen metabolism is accelerated with 0.4 μmol·L^(-1) NdCl_3 treatment.

Effects of Lanthanum on Root Growth and Senescence of GF_(43) (Prunus domestica) Plantlet in Vitro

The effect and the action mechanism of lanthanum on GF_(43) plantlet in vitro were studied. The results of experiments show that root growth rate and dry weight of GF_(43) by LaCl_3 treatments increase. The activities of antioxidant enzymes in root system such as superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD) apparently enhance. Moreover ·O^-_2 and malond ialdehyde (MDA) contents and cell membrane permeability of GF_(43) are decreased by LaCl_3. The relatively stable membrane structure of cell could also be maintained and the root ageing of GF_(43) plantlet in vitro delays.

Effect of Lanthanum on Deferring Root Ageing of Peach Plantlet in Vitro

Poor adventitious root formation is a major obstacle in micropropagation and in conventional propagation. The effects of LaCl_3 on root growth and deferring the root ageing of peach plantlet in vitro were investigated. The results show that the optimum concentration of LaCl_3 (2.5 μmol·L -1) in the rooted medium can significantly increase the rooting rate, the root length and the fresh weight, promote the activities of superoxide dismutase (SOD); catalase (CAT) and peroxidase (POD) and decrease the production rate of O_2 ·-, the malondialdehyde (MDA) contents and the plasma membrane permeability. There are important theory meaning and practical value in applying LaCl_3 in the rooting medium to raise the rate of rooting and transplant for stone fruit tree.

REGULATION OF PHYTOHORMONS ON THE REGENERATION OF SHOOT-TIPS OF ANTIRRHINUM MAJUS INTO PLANTLET IN VITRO

Tissue culture of Antirrhium majus was experimented by using the tip of shoots as the explants, and comparing the effects of NAA, IAA and 2, 4-D on the formation of adventitious buds. The results indicated that the effect of NAA was the best for the formation of adventitious buds, 5.0, 7.0, 9.0, 11.0, 13.0 mg/L BA combinatcd with 0.1, 0.5, 1.0, 1.5, 2.0 mg/L NAA respectively. The results show that combinations of the concentrations of BA 7.0-9.0 mg/L and NAA 0.1-0.5 mg/L were advantageous to the regeneration of buds and NAA was very important for callus growth.