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 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.

Construction and Characterization of cDNA Library from Water-Stressed Plantlets Regenerated in vitro of Populus hopeiensis

In order to isolate and clone water-stress-responsive genes, total RNA was extracted from water-stressed plantlets regenerated in vitro of Populus hopeiensis using a QIAGEN RNeasy Plant Mini Kit. CDNA, synthesized by LD-PCR with the SMART cDNA Library Construction Kit, was in vitro packaged into a phage λTriplEx2 vector. The resulting primary library and amplified library have a titer of 1.68×10^6 and 1.69×10^9 pfu·mL^-1 respectively. The combination ratio reached 98.8% and the average size of inserts was about 800 bp. In addition, the percentage of inserted fragments （〉400bp） was approximately 90%. The results indicate that a cDNA library has been successfully constructed.

Effects of Sion Cold Resistance of Cymbidium Plantletsin vitro

To determine the medium with the concentration of Na2SiO3 , the effect of different concentrations of Na2SiO3 on cold resistance of Cymbidium plantlets in vitro was studied. The results showed that Si enhanced osmotic regulation ability of Cymbidium evidently on cold resistance with increasing contents of free Pro, soluble sugar, soluble protein and decreasing contents of MDA. Compared with CK, osmotic regulation substances are accumulated rapidly. Free Pro, soluble sugar, soluble protein, MDA could be adopted as reference indices to evaluate capability of frost resistance of Cymbidium. By experiment we stated Na2SiO3 (3mmol/L) could relieve low temperature injury of Cymbidium plantlets in vitro. Mitigative effect was reduced with increasing concentration of Na2SiO3 .

Generation of Virus Free Potato Plantlets through Meristem Culture and Their Field Evaluation

Different aspects of micropropagation through meristem culture for the production of virus indexed source plants, in vitro tuberization and field evaluation of the in vitro regenerated plants were studied on four commercial cultivars of potato (Solanum tuberosum L.) viz., Diamant, Cardinal, Shilbilati and Lalpakri. The investigation was conducted at Rajshahi, Bangladesh from December 2010 to March 2012 to produce virus-free potato plantlets through meristem culture, shoot multiplications with root induction as well as their acclimatization and evaluation of morphological characters and tuber yield under field condition. Shoot tips of 25 - 30 day old field-grown plants of above mentioned four cultivars were used for meristem isolation. After isolation, meristems of these varieties of potato were cultured on “M” shaped filter paper bridge in Murashige and Skoog (MS) liquid medium. Four different treatments of media formulations viz. 0.1 mg/L KIN + 0.1 mg/L GA3, 0.1 mg/L KIN + 0.5 mg/L GA3, 0.5 mg/L KIN + 0.1 mg/L GA3 and 0.5 mg/L KIN + 0.5 mg/L GA3 were used as plant growth regulators. From these formulations MS + 0.1 mg/L KIN + 0.5 mg/L GA3 was found to be the best for the primary establishment of meristem culture. The primarily established meristems were subcultured on to MS semisolid basal medium supplemented with four different treatment combinations of hormones viz. 0.5 mg/L BA + 1.0 mg/L IBA;0.1 mg/L KIN + 0.1 mg/L GA3;0.5 mg/L BA + 0.5 mg/L GA3 and 0.5 mg/L KIN + 0.5 mg/L GA3 were used to identify the suitable media compositions for shoot proliferation. Results showed that out of these four media treatments the formulation 0.5 mg/L BA + 0.5 mg/L GA3 was found to be the best suitable for shoot generation. Among the four cultivars of potato higher frequency of shoot proliferation (number of shoots/explant and longest shoot length) was observed in Diamant, though the highest shoot formation (76%) was recorded in Cardinal. Virus free in vitro grown potato plantlets were obtained through DAS-ELISA test and used substantially for micro-propagation. After gradual acclimatization of rooted plantlets of four potato cultivars, they were transferred into the field for cultivation and established successfully. It was observed from the field study of in vitro meristem-derived plantlets that there were no virus-affected plants. The virus-free exotic varieties were much superior in all vegetative attributes and yield compared to those of indigenous varieties with producing potato plants of normal height. In contrast, the indigenous varieties took a longer time to tuber initiation and maturity, lower plant height and number of leaves per plant, a higher number of tubers but a lower amount of tuber weight per plant, and poorer tuber grade than the exotic varieties. However, the variety Cardinal exposed the best performances in the context of survival percentage of plantlets (90%), days to tuber initiation (DTI), a number of leaves per plant (NL), tuber weight per plant (343.40%) and the percentage of rich tuber grade.

Study on Efficient Multiplication of Blueberry(Vaccinium corymbosum)Plantlets in Swinging type Temporary Immersion Bioreactors System(S-TIBs)

The swinging type temporary immersion bioreactors system （S-TTBs） is a kind of new and advanced method of tissue culture. The efficient multiplica- tion technology of blueberry （ Vaccinium corymbosum） plantlets in S-TTBs was systematically studied through L9 （3^4） orthogonal experiment with three factors （culture medimn volmne, swing angle, and inoculation density） and completely randomized experiment with two factors （swing frequency and immersion time）. The results showed that the optimal culture parameters were set as follows： the culture medium volume of 250 nil/bottle, the swing angle at 45 ° , and the inoculation density of 60 plantlets/bottle. The optimal swing frequency was 1 time/6 h, and the immersion time was set as 60 s.

Effects of Non-Conventional Substrate and NPK Fertilizer on Cassava Plantlets Acclimatization for Plant Material Production

The adaptation of cassava plantlets to natural conditions remains the main obstacle in the process of free virus plantlets production. In this study, cassava plantlets acclimatization was assessed by pre-acclimatization, height of acclimated plantlets, substrate, and nutrient watering solution. The results revealed that pre-acclimatization gave a high survival rate (61.9%) whereas the direct tunnel acclimatization was conducted to low rate (27.31%) of plantlets. Furthermore, the height of plantlets influenced significantly (p < 0.05) their survival rate with best survival rate observed on plantlet height from 1 cm to 5 cm. There was no significant difference between the substrates used and watering solutions. However, the Sawdust substrate gave a high survival rate (46.67%) and better phylogenesis (1.8) in the variety RB 89509. Likewise, plantlets treated with NPK nutrient watering solutions (20-10-10) had a better survival rate (48.57%), better phylogenesis (2.125) with the variety RB 89509 and better growth in height (1.18 cm) with the variety BF92/0267. These findings constitute a database for the establishment of the technical itinerary for the acclimatization of cassava plantlets.

2-20 Synthesis of Uniform Sizes of Uranium Dioxide Kernels by Test-tube Experimrent in the Laboratory

In the 1970s and 1980s, United States Department of Energy conducted numerous studies on the fabrication of nuclear fuel particles using the internal gelation process[1], the internal gelation process is an advanced production process for nuclear fuels. Recently, we had used these lessons learned on the internal gelation process to prepare quantities of uranium dioxide kernels of uniform sizes. This paper discusses the whole efforts of experimental processes.

Influence of Watering Solution and Phenotype on the Growth of <i>in Vitro</i>Propagated Pineapple (Smooth Cayenne Cultivar) Plantlets during Acclimatization

Pineapple is the first fruit crop cultivated in south Benin that greatly contributes for food and nutritional security and farmers’ income. But the lack of homogenous planting material constitutes the major constraint for improving pineapple yield. In vitro micropropagation is now used in the production of homogenous and free disease planting materials of pineapple. However, the acclimatization to natural condition of pineapple plantlets is an important step in planting material production of this crop. Here, we determined the intrinsic and extrinsic factors which influence the behavior of plantlets during the acclimatization process. For this purpose, plantlets from different categories were selected, trimmed and planted on a horticultural substrate made up of potting soil, white sawdust and compost previously sterilized. The plantlets were then incubated in under acclimatization greenhouse with average temperature of 29˚C and 70.2% of humidity. A batch of plantlets was subjected to two different watering solutions: Shive and Robbins solution and NPK 14-6-5 foliar fertilizer. The results obtained initially showed high rate (100%) of survival and growth of the plantlets watered with Shive and Robbins solution against 50% of the plantlets watered with the foliar fertilizer solution. In addition, the plantlets with spread pores exhibiting the characteristics of which the number of leaves varies between 9 and 11, the weight between 1.2 and 1.5 g, the size of 4.5 to 5.5 cm, and a good junction between the aerial part and the root system were those which were successfully grown in acclimatization phase under greenhouse, unlike plantlets with erected pores having lower success rate. This study goes a long way in providing good procedures of acclimatization of homogenous and free disease planting material of pineapple to the famers.

Effect of desiccation on regeneration frequency of green plantlets and some biochemical properties of calli of wild rice

Wild rice is an important resource of usefulgenes to rice breeders. However, low regener-ation frequency is an obstacle to use the valu-able genes. We used desiccation to improve theregeneration frequency and studied the bio-chemical changes of calli of wild rice after des-iccation.Materials used in this experiment werewild species O. rufipogon, O. meyeriana, O.alta, and O. brachyantha. Young panicles(0.1-0. 5 cm in length of the inflorescence)

碳离子束辐照对马铃薯组培苗生长及生理特性的影响

为研究高能碳离子束对马铃薯组培苗茎切段的影响,以马铃薯品种‘新大坪’的组培苗茎切段为研究材料,设置4个辐照剂量梯度对其进行辐照处理,测定植株致死率、生根率、侧芽形成率、形态变异率以及叶片生理指标,确定‘新大坪’组培苗茎切段的半致死辐照剂量。结果表明:辐照剂量与致死率之间的关系呈正相关性;组培苗茎切段的再生植株表型变化主要为叶片不对称、叶缘不规则、茎变异伸长、叶片丛生和叶片粘连等;随辐照剂量升高,超氧化物歧化酶、过氧化物酶和过氧化氢酶活性呈先升后降的趋势;丙二醛含量则呈先升后降,随后又升高的现象。利用线性回归方程计算‘新大坪’的半致死辐照剂量为20.37 Gy。

珙桐体细胞胚胎发生技术研究

研究探讨基本培养基类型对珙桐愈伤组织诱导的影响、植物生长调节剂对胚性愈伤组织诱导及体胚发育的影响,初步建立以未成熟合子胚为外植体,通过体细胞胚胎发生途径再生珙桐植株的培养方案。结果表明,在含有3%蔗糖、0.3%凝胶、800 mg/L水解酪蛋白、400 mg/L L-谷氨酰胺的MS、1/2MS和WPM基本培养基上均可诱导出愈伤组织,以MS+2.0 mg/L 2,4-D+0.5 mg/L 6-BA诱导率最高,可达94.0%;胚性愈伤组织在MS+0.5 mg/L NAA+1.0 mg/L 6-BA的培养基上诱导效果较好;在MS+0.5 mg/L NAA+1.0 mg/L 6-BA培养基上,体胚诱导率可达22.9%;成熟体胚在1/2MS+0.5 mg/L 6-BA+0.25 mg/L IBA的萌发培养基上能正常萌发,其后在添加1 g/L活性炭的萌发培养基上转化成完整小植株。组织细胞学观察表明,珙桐体胚起源于胚性愈伤组织,经历了球形期、心形期、鱼雷期和子叶期发育阶段,这与自然条件下的合子胚形成过程相似。

云南马铃薯不同品种(系)对马铃薯疮痂病的抗性分析

随着云南省马铃薯种植面积的扩大和种薯调运的频繁,质量监管不足导致马铃薯疮痂病的发生日益严重。本研究旨在评估云南省马铃薯主栽品种对该病害的抗性水平,并筛选出具有潜在抗病性的育种材料。为此,本研究收集了33份不同的马铃薯品种(系)材料,并通过组织培养方法培育组培苗,进而采用盆栽试验进行抗性评估。同时结合对大田主栽品种田间发病情况的调查。结果显示,供试的33份马铃薯品种(系)的组培苗接种疮痂病菌后,所有测试品种均出现了不同程度的病害,发病率58%~96.67%,其中I-1085品种的发病率最低,F37-2品种的发病率最高。病情指数分布在22.58~67.27之间,以Fx3品种的病情指数最低,F37-2的最高。本研究结果为云南省马铃薯抗疮痂病品种选育工作提供了重要的基础性数据和理论支撑。

氮缺乏对84K杨树组培苗光合特性及次生代谢的影响

研究氮缺乏对84K杨树组培苗生长的影响,以期为84K杨的育种与繁殖提供理论基础。以84K杨树组培苗为实验材料,通过氮缺乏处理,测定84K杨树组培苗的生物量、光合参数、叶绿素荧光参数,并进一步分析氮缺乏时全氮、全碳、碳氮比、总酚及抗氧化活性的变化。结果显示:随着培养基中氮含量的减少,84K杨树组培苗的干重、净光合速率(P n)和气孔导度(G s)均受到抑制,同时,氮缺乏导致84K杨树组培苗叶绿素a、叶绿素b、总叶绿素含量及最大光化学效率(Fv/Fm)降低。此外,随着培养基中氮素的减少,84K杨树组培苗中全氮含量显著减少,碳氮比显著升高,总酚含量显著增加。抗氧化分析结果表明,培养基中氮含量越低,组培苗的抗氧化活性越好。相关性分析表明,生物量与光合参数及叶绿素含量之间存在极显著正相关,但与总酚含量之间存在负相关;全氮含量与总酚含量之间存在负相关,与全碳及碳氮比之间存在正相关;同时84K杨树组培苗总酚含量及抗氧化活性之间具有明显的相关性。研究结果表明:氮缺乏可对84K杨树组培苗的光合生理及次生代谢产物产生影响,不利于84K杨树组培苗的生长和光合作用,但促进了多酚等抗氧化物质的产生。本研究可为84K杨树的栽培管理提供理论依据和基础数据。

金钗石斛组培苗壮苗培养研究

为研究金钗石斛组培苗快速生长的影响因素,以生长素NAA、水解乳蛋白、MS培养基有机成分、活性炭四因素三水平设置正交试验,观察金钗石斛组培苗壮苗培养的生长表现,从而确定最佳培养基组合。试验结果表明:0.10 g/L活性炭+0.50 mg/L NAA+0.05 g/L水解乳蛋白+1倍有机成分组合对组培苗各生长效果最好;加入NAA可以明显增加组培苗生长的高度和茎粗。本研究可为金钗石斛组培快繁研究奠定基础。