Transcriptome Analysis of Inflorescence Development at the Five-Leaf Stage in Castor(Ricinus communis L.)

The yield of castor is influenced by the type of inflorescence and the proportion of female flowers.However,there are few studies on the genetic mechanism involved in the development and differentiation of castor inflorescences.In this study,we performed transcriptomic analyses of three different phenotypes of inflorescences at the five-leaf stage.In comparison to the MI(complete pistil without willow leaves),290 and 89 differentially expressed genes(DEGs)were found in the SFI(complete pistil with willow leaves)and the BI(monoecious inflorescence),respectively.Among the DEGs,104 and 88 were upregulated in the SFI and BI,respectively,compared to the MI.In addition,186 DEGs and 1 DEG were downregulated in the SFI and BI compared to the MI.Moreover,we conducted GO and KEGG enrichment analyses of the DEGs.In comparison to the MI,the SFI and BI exhibited the enrichment of functional branches in DEGs,specifically in pollen wall assembly,pollen development,and cellular component assembly involved in morphogenesis.In our study,RADL5 showed low expression levels between SFI-vs.-MI types.In addition,we found that the expression of NAC in the SFI differed from that in MI and BI,and some genes related to hormonal signaling changed their expression levels during inflorescence differentiation.These results reveal the genetic mechanism of sex genotypes in castor,which will not only guide researchers in the breeding of castor but also provide a reference for genetic research on other flowering plants.

Proteomic Analyses of Three Inflorescence Styles of Castor(Ricinus communis L.)at Different Developmental Stages

Castor(Ricinus communis L.)is one of ten oil crops in the world and has complex inflorescence styles.Generally,castor has three inflorescence types:single female inflorescence(SiFF),standard female inflorescence(StFF)and bisexual inflorescence(BF).StFF is realized as a restorer line and as a maintainer line,which was applied to castor hybrid breeding.However,the developmental mechanism of the three inflorescences is not clear.Therefore,we used proteomic techniques to analyze different inflorescence styles.A total of 72 diferentially abundant protein species(DAPs)were detected.These DAPs are primarily involved in carbon and energy metabolism and carbon fixation in the photosynthetic organism pathway.The results showed that DAPs are involved in photosynthesis to control the distribution of imported carbohydrates and exported photoassimilates and thus affect the inflorescence development of castor.In addition,these DAPs are also involved in cysteine and methionine metabolism.Quantitative real-time PCR(qRT-PCR)results demonstrated that the proteomics data collected in this study were reliable.Our findings indicate that the carbon cycle and amino acid metabolism influence the inflorescence development of castor.

<i>Wrinkled</i>1 (WRI1) Homologs, AP2-Type Transcription Factors Involving Master Regulation of Seed Storage Oil Synthesis in Castor Bean (<i>Ricinus communis</i>L.)

Among APETALA2 (AP2)-type plant specific transcription factor family, WRINKLED1 (WRI1), has appeared to be a master gene transcriptionally regulating a set of carbon metabolism- and fatty acid synthesis (FAS)-related genes responsible for seed specific triacylglycerols (TAGs) storage in oil plants. B3 type transcription factors, such as ABI3 and FUS3, are known to be involved in seed development, such as seed storage protein synthesis and maturation. Based on the recent whole genome sequence data of castor bean (Ricinus communis L.), putative WRI1 homologs (RcWRI1, RcWRI2) specifically expressed in castor bean seed have been identified by comparing organ specific expression profiles among seed development-related transcription factors, seed storage specific genes (Ricin, RcOleosin) and a set of FAS genes including genes for sucrose synthase (RcSUS2), biotin carboxyl carrier protein (a subunit of acetyl-CoA carboxylase, RcBCCP2) and ketoacyl-acyl carrier protein synthase (RcKAS1). Immunoreactive signals with WRI1, FUS3 and ABI5-related polypeptides were also detected in seed specifically, consistent with the expression profiles of seed development-related genes. The WRI1 binding consensus sites, [CnTnG](n)(7)[CG], designated as the AW-box, were found at the promoter region of RcBCCP2 and RcKAS1. Thus, RcWRI1 possibly play a pivotal role in seed specific TAGs storage during seed development by directly activating FAS -related genes.

Genome-wide Identification,Classification and Expression Analysis of Lhc Supergene Family in Castor Bean(Ricinus communis L.)

Light-harvesting chlorophyll a/b-binding （LHC） proteins are a group of nuclear-encoded thylakoid proteins that play a key role in plant photosynthesis and are widely involved in light harvesting, energy transfer to the reaction center, maintenance of thylakoid membrane structure, photoprotection and response to en- vironmental conditions, etc. Although/dw supergene family is well characterized in model plants such as Arabidopsis, rice and poplar, little information is available in castor bean （Ricinus communis L. ）. In this study, a genome-wide search was carried out for the first time to identify castor bean L/w genes and analyze the gene structures, biochemical properties, evolutionary relationships and expression characteristics based on the published data of castor bean genome and ESTs. According to the results, a total of 28 Rclhcs genes representing 13 gene families （ l_hca , l_hcb , Elip , Ohpl , Ohp2 , SEP1, SEP2 , SEP3 , SEP4 , SEP5 , PsbS , Rieske and FCII） and 25 subgene families were identified in castor bean genome; to be specific, 25 and 5 genes were found to have corresponding ESTs in NCBI and have al- ternative splicing isoforlns, respectively. These RcLhcs contain 0 to 9 introns and distribute on 26 of the 25 878 released scaffolds. All RcLhcs genes were found to be expressed in all examined tissues, i.e. leaf, flower, II/III stage endosperm, V/VI stage endosperm and seed, with the highest expression level in leaf tissue.

In vitro establishment of a highly effective method of castor bean (Ricinus communis L.) regeneration using shoot explants

An efficient plant regeneration protocol was established for castor bean （Ricinus communis L.）, in which 0.3 mg L-1 thidiazuron （TDZ） induced shoot clusters and increased the number of adventitious shoots from hypocotyl tissue. Our results showed that treatment under dark conditions significantly promoted the average number of shoots per explant to 37.36±4.54 （with a 6-d treatment）. Modified 1/2 Murashige and Skoog （MS） basal medium supplemented with 440 mg L-1 Ca2＋, 0.2 mg L-1 gibberellic acid and 0.1 mg L-1 TDZ significantly increased shoot elongation rates and lowered vitrification rates. Further- more, 1/2 MS media supplemented with 0.2 mg L-11-naphthaleneacetic acid induced a higher rooting rate compared with other culture conditions.

Study on Cu Resistance of Ricinus communis L. Callus

[Objective] The article studies the growth and Cu absorption of Ricinus communis L. callus under Cu stress. [Method] CuSO4.5H20 solutions with different Cu concentrations were added to callus subculture medium; callus was inoculated and Cu resistance index of callus was worked out. Cu content in callus was deter- mined with the method of Varian AA240FS. [Result] With the Cu concentration at 60 mg/L, the growth of callus was inhibited, its Cu resistance index was only 33.87%. With the Cu concentration at 40 mg/L, callus was faint yellow in color, and grew rapidly with its Cu resistance index at 61.29%. Such high level resistance could remain the same after six week after continuous subculture. In the 4t week of culture, Cu resistance index in treatments with Cu concentrations at 10, 20, 30, 40 mg/L was higher than that in the 3rd week, and the content of Cu in callus of the treatments was 0.33, 0.54, 1.16, 1.40 mg/g respectively. [Conclusion] Cu con- centration at 40 mg/L in culture medium can be the threshold for selecting Cu re- sistance R. communis callus.

Studies on Cell Suspension Culture of Ricinus communis L.

Ricinus communis L. is a new copper hyperaccumulator growing on Tonglushan copper mine in Hubei Province, China. This study aimed to establish a suspension cell line of R. communis L. with stable and rapid growth for further screening of heavy metal-resistant R. communis L. cells and breeding of hyperaccu- mulators. In this study, cell suspension culture conditions were optimized by using orthogonal experimental design with previously induced R. communis L. embyre- genic calluses as experimental materials, to establish the suspension cell line of R. communis L. Under the optimal conditions, growth curves of suspension cells and changes in pH of culture liquid were determined. The results showed that the optimal culture conditions for R. conmmnis L. suspension cell line were ： MS ＋ O. 5 rag/I, 6-BA ＋ O. 2 mg/L NAA ＋ 50 mg/L sucrose ＋ 350 mg/L casein hydrolysate as basic medium, with callus inoculation amount of 2.5 g/50 ml, dark culture at （26 ±2） ℃ with shaking at 110 r/min. Under these conditions, increment of fresh weight and dry weight ofR. commun/s L. suspension cells reached the maximum of 4.56 g/（50 ml 14 d） and 0.49 g/（50 ml 16 d）, respectively. Growth curves of R. communis L. suspension cells were basically in ＂S＂ shape. Each culture cycle lasted 16 d, and the rapid growth stage was from the 6th d to the 14th d. In a culture cycle, pH of the culture liquid declined first and then increased to the maximum and stabilized gradually.

Silencing of the Nonspecific Phospholipase C6(NPC6)Gene Induces Ricinoleic Acid Accumulation in Castor Seeds

Castor,scientifically known as Ricinus communis L.,is among the top ten oil crops globally.It is considered a renewable resource and is commonly referred to as‘green oil’.Castor seeds contain castor oil as their main component,which is predominantly composed of ricinoleic acid.This study utilized RNAi technology to silence the NPC6 gene in NO.2129 castor,resulting in the creation of mutant plants L1 and L2.The weight of 100 dry seed kernels from L1 and L2 exceeds that from NO.2129.The crude fat and ricinoleic acid levels of L1 and L2 were higher than those of NO.2129 at various developmental stages.In the proteomics analysis of 60-day-old castor seeds,a total of 21 differentially expressed proteins were identified,out of which 19 were successfully recognized.Eleven of the differentially expressed proteins identified were legumins,which play a crucial role in nutrient storage within the seed.Silencing the NPC6 gene results in the accumulation of ricinoleic acid in castor seeds.The findings of this study not only enhance our knowledge of NPC6’s role in regulating castor seed oil synthesis but also offer fresh perspectives for investigating oil synthesis and accumulation in other plant species.

Intraspecific DNA methylation polymorphism in the non-edible oilseed plant castor bean

Investigation of the relationships of phenotypic and epigenetic variations might he a good way to dissect the genetic or molecular basis of phenotypic variation and plasticity in plants, Castor bean （Ricinus cornraunis L）, an important non-edible oilseed crop, is a mono-species genus plant in the family Euphorbiaceae. Since it displays rich phenotypic variations with low genetic diversity, castor bean is a good model to investigate the molecular basis of phenotypic and epigenetic variations. Cytosine DNA methylation represents a major molecular mechanism of epigenetic occurrence. In this study, epigenetic diversity of sixty landrace accessions collected worldwide was investigated using the methylation- sensitive amplification polymorphism （MSAP） technique, Results showed that the epigenetic diversity （based on the polymorphism of DNA methylated loci） exhibited a medium variation （Ne = 1.395, He = 0.242, I = 0.366） at the population level though the variation was great, ranging from 3,80% to 3431% among accessions. Both population structure analysis and the phylogenetic construction （using the neighbor-joining criteria） revealed that the two main clades were identified, but they did not display a distinct geographic structure, After inspecting the location of polymorphic methylated loci on genome we identified that the polymorphic methylated loci occur widely in nuclear and organelle genomes. This study provides new data to understand phenotypic and epigenetic variations in castor bean,

Integration of morpho-physico-biochemical traits with SSR and SRAP markers for characterization of castor genotypes of Indian origin

Castor(Ricinus communis L.)is an important tropical oilseed crop,whose oil has versatile,practical value,especially in industries.The present study aimed to estimate the nature and magnitude of variability in the castor germplasm concerning yield and its component traits and physico-biochemical characters.Seed yield per plant and oil content ranged from 80.90 g(ICS-165)to 248.30 g(RG-3216),and 34.7%(ICS-172)to 58.7%(JI-277),respectively.The iodine value of oil ranged from 76.36(JI-370)to 89.84(P2-135)with an average value of 83.02.The mean saponification value of oil was 182.24.The genotypic and phenotypic coefficients of variation were high for acid value,capsules on the main raceme,seed yield per plant,and total length of the main raceme.A positive association of porosity,average unit volume,and total length of the main raceme with seed yield per plant showed that these characters might be directly attributed to seed yield improvement.By Manhattan distances,the 30 genotypes were grouped into 3 clusters.Their genetic diversity was elucidated using SSR and SRAP markers.SRAP marker produced higher mean number of total bands(5.71),polymorphic bands(4.57),percentage polymorphism(83.10%),PIC(1.72),RP(5.90),mean RP(1.02),MR(5.71),EMR(4.57)and MI(1.44)values when compared to SSR(2.89,2.11,79.63%,0.61,1.90,0.72,2.89,2.11 and 0.49,respectively)marker.The highest genetic distance(0.77)was between 48-1 and JI-370,which indicated that these genotypes could be used in biparental mating schemes,QTL map development,and hybridization programmes to increase oil content and quality for industrial purposes.

Response of Castor Bean Plants to Different Row Spacings and Planting Seasons

The cultivation of the castor bean plant is prominent and important because of its potential for producing vegetable oils with special properties. However, research on cultivating castor bean plants is scarce, especially in areas with altitudes of 300 m or below. Moreover, the currently available cultivars have not been widely tested in Brazil. Thus, the aim of this study was to evaluate the response of the castor bean cultivar BRS Nordestina to different spacings at low altitudes during two cropping seasons. The study was conducted in the municipality of Itaocara, in the northwest Fluminense region, Brazil, which has an altitude of 60 m. The treatments consisted of five row spacings (1.5, 2.0, 2.5, 3.0 and 3.5 m) with 1 m between plants. The plants were cultivated in the 2007/2008 spring/summer season or in the 2008 autumn/winter season. A randomised block design with four replicates was used. The following parameters were assessed: plant height, height of insertion of primary raceme, number of racemes per plant, raceme length, number of fruits per raceme and grain yield. The spacing of 2.5 m between rows had the highest yield of castor beans during the two cropping seasons. The average yield of the cultivar BRS Nordestina indicates its potential for cultivation in northern Rio de Janeiro, below an altitude of 300 m. The yield was higher during the summer cropping season. The less-dense arrangement of the plant population contributed to the cultivation of plants with more racemes in both growing seasons.

Cytocompatible cellulose nanofibers from invasive plant species Agave americana L.and Ricinus communis L.:a renewable green source of highly crystalline nanocellulose

In this study,the fibers of invasive species Agave americana L.and Ricinus communis L.were successfully used for the first time as new sources to produce cytocompatible and highly crystalline cellulose nanofibers.Cellulose nanofibers were obtained by two methods,based on either alkaline or acid hydrolysis.The morphology,chemical composition,and crystallinity of the obtained materials were characterized by scanning electron microscopy(SEM)together with energy-dispersive X-ray spectroscopy(EDX),dynamic light scattering(DLS),X-ray diffraction(XRD),and Fourier transform infrared(FTIR)spectroscopy.The crystallinity indexes(CIs)of the cellulose nanofibers extracted from A.americana and R.communis were very high(94.1%and 92.7%,respectively).Biological studies evaluating the cytotoxic effects of the prepared cellulose nanofibers on human embryonic kidney 293 T(HEK293 T)cells were also performed.The nanofibers obtained using the two different extraction methods were all shown to be cytocompatible in the concentration range assayed(i.e.,0-500μg/mL).Our results showed that the nanocellulose extracted from A.americana and R.communis fibers has high potential as a new renewable green source of highly crystalline cellulose-based cytocompatible nanomaterials for biomedical applications.

蓖麻高产宜机收杂交组合评价

【目的】对蓖麻杂交组合进行联合评价,为筛选高产、宜机收蓖麻品种提供理论参考。【方法】联用灰色关联度分析、主成分分析和聚类分析对30个蓖麻杂交组合的产量性状、宜机收性状和光合性能3个方面进行综合评价。【结果】在灰色关联度分析中,根据育种目标构建了理想品种;通过各组合与理想品种之间的加权关联度评选出N_(19)(0.768)、N_(7)(0.751)、N_(11)(0.727)、N_(10)(0.717)、N_(6)(0.713)和N_(13)(0.712)6个组合。其中,前4个组合的加权关联度优于淄蓖5号(0.714),后2个组合的加权关联度与淄蓖5号接近;主成分分析结果将10个性状归纳为株型因子、产量决定因子和光合与分枝夹角因子3个主成分,累计贡献率达67.07%。30个组合中,21个组合的综合得分均大于淄蓖5号(-0.699),且仅N_(18)(2.370)、N_(4)(1.848)、N_(19)(1.742)和N_(11)(1.019)4个组合的得分大于1,并以N19最为突出,其得分远高于其他材料;聚类分析结果显示,在欧氏距离值为22处可将30个组合划分为4个类群,第Ⅳ类群成员(包括N19、N11和N30)的整体表现与理想品种最为相似,但分枝夹角(59.10°)较大,仍需进一步改良。【结论】综合3种评价结果,避免了多余材料入选和优良材料丢失,最终筛选出N19和N112个产量高、株型好和光合速率高的蓖麻杂交组合,它们的综合表现均优于淄蓖5号,为品种改良方向提供参考。

蓖麻LEA基因家族的全基因组鉴定、分类及其进化分析

基于已公布的基因组和EST数据对蓖麻LEA基因家族进行全面鉴定和系统命名，并在此基础上分析了其基因结构、生化特征、进化关系和表达特性。结果显示，蓖麻基因组中存在27个LEA基因，分属于LEA_1、LEA_2、LEA_3、LEA_4、LEA_5、LEA_6、dehydrin和SMP 8个亚家族，它们散布于24条scaffold上，含有0～2个内含子不等，且其中有3个基因存在可变剪接；根据编码蛋白中的Pfam结构域类型及其进化关系，将这些LEA基因依次命名为RcLEA1-1和-2、RcLEA2-1和-2、RcLEA3-1至-3、RcLEA4-1至-7、RcLEA5-1和-2、RcLEA6-1和-2、RcLEA7-1至-5和RcLEA8-1至-4；利用BLAT分析基因表达谱显示，在叶片、花、II/III期胚乳、V/VI期胚乳和种子等组织中，所有RcLEA基因都有表达；启动子分析表明，RcLEA基因的启动子区富含LTRE、ABRE、MYC、MYB和W-box等逆境应答相关顺式作用元件。

蓖麻RcNFYA3-1基因的克隆及表达分析

为探讨NFYA3-1基因在蓖麻耐盐中的作用,以哲蓖三号植株为材料,对RcNFYA3-1进行克隆,获得完整的编码区序列(CDS),并对其理化性质和同源性等进行预测分析,利用qRT-PCR分析RcNFYA3-1基因在盐胁迫下的表达。结果发现:RcNFYA3-1基因全长1565 bp,CDS全长984 bp,编码327个氨基酸的不稳定疏水蛋白,预测其蛋白质分子量为36.093 kD,不存在信号肽;其二级以无规则卷曲为主。Rc-NFYA3-1蛋白与木薯和巴西橡胶树的同源性最高。qRT-PCR分析结果表明,RcNFYA3-1基因在蓖麻各部位均有表达,且主要在根中发挥作用。

蓖麻基因组中一种Mutator-like类型MITEs元件的结构分析和鉴定

利用生物信息学方法,对蓖麻基因组Mutator-like类型MITEs元件进行了分析和鉴定。结果表明,蓖麻基因组上共鉴定出76个小于3 kb的Mutator-like类型MITEs元件,其中长度小于1 000 bp的元件有53个,占69.7%。这些MITEs元件中有42个位于基因间隔区,11个位于基因内含子区,位于5'UTR和3'UTR区的分别有8个和4个,另有1个位于基因编码区。29个MITEs元件带有转座酶以外的功能基因片段,成为pack-MITEs。多个pack-MITEs带有相同的非转座酶基因片段。Mutator-like类型的MITEs对蓖麻基因的表达和基因组的进化有潜在的影响,值得深入分析。

蓖麻油体蛋白基因家族鉴定和盐碱胁迫响应分析

本研究以蓖麻为对象,利用生物信息学手段对蓖麻Oleosin基因家族进行全面鉴定分析,得到5个Oleosin基因家族成员,序列长度在138~232 aa之间;分子量在14.48493~25.42461 kDa之间;等电点介于9.07~10.96之间;均为疏水性蛋白且具有高度保守的“脯氨酸结”;α-螺旋和无规则卷曲是蓖麻Oleosin蛋白的主要组成结构;基序分析显示,Motif1~Motif4存在所有蓖麻Oleosin蛋白中。编码这些蛋白的5个Oleosin基因分布于4条染色体上,在进化关系上被划分到U-Oleosin、SL-Oleosin和SH-Oleosin三个亚族;基因结构分析显示,除RcOleosin2有一个内含子外,其余RcOleosin基因均无内含子;5个基因启动子区域均存在多种与逆境响应相关的顺式作用元件;共线性分析显示,基因组多倍化不是蓖麻Oleosin基因家族扩张的主要原因;盐碱胁迫下的蓖麻种子转录组数据表明,5个Oleosin基因均在盐碱胁迫过程中有一定的调节功能。

蓖麻MYB转录因子家族的鉴定和生物信息学分析

MYB是植物中最大的转录因子家族之一,在植物生长发育和逆境胁迫响应等多个生理生化过程中具有重要作用,但目前缺乏对蓖麻的MYB转录因子的系统分析.首先基于基因组挖掘出32个转录因子,并对其进行蛋白基序、功能注释、系统进化以及理化性质的分类鉴定;其次基于转录组数据分析了胁迫条件下这些转录因子的表达谱,分析其对胁迫条件的响应.结果表明蓖麻在盐胁迫下和热胁迫下MYB蛋白表达量蓖麻的生长有影响.研究结果为进一步研究MYB转录因子在蓖麻非生物胁迫中的功能提供参考.

蓖麻枯萎病抗性的QTL定位分析

本研究用YC2×YF1抗、感组合的F2群体在两个环境中对蓖麻枯萎病抗性进行了QTL定位。遗传分析表明,群体中枯萎病抗性呈连续性偏态分布,后代表型偏抗性亲本。2014年检测到5个QTL,解释了总变异的18. 21%,单位点贡献率为0. 05%～12. 32%。2015年分别在出苗后20d、30d、40d、现蕾期和乳熟期进行了动态定位,分别检测出4、12、13、4和1个QTL,单位点贡献率为1. 64%～21. 91%,5个发育时期检出的QTL主要集中在第3、4和8连锁群上。有1个QTL在4个时期、3个QTL在3个时期、6个QTL在2个时期被重复检测到。在第8连锁群上一个QTL在两个环境同一时期被重复检测到。以上结果可为蓖麻抗枯萎病分子标记辅助选择提供参考。

蓖麻内源防御肽的MALDI-MSI方法及溯源应用研究

国际范围内,蓖麻毒素、蓖麻籽粗提物以及蓖麻籽等均为蓖麻毒素信件的潜在来源,屡屡引发公共安全威胁。因此,蓖麻籽产地溯源至关重要。该研究选择高丰度、低毒性的蓖麻内源性防御肽(RCB),系统探讨优化了冷冻切片制备、基质喷雾等条件,首次建立了RCB的基质辅助激光解吸电离-质谱成像(MALDI-MSI)方法。选择四川、山西为中国南北代表性产地,阐述蓖麻籽组织切片中3种RCB(RCB 1、RCB 2、RCB 3)的丰度及空间分布特点。结果表明,3种RCB主要分布在成熟种子的胚乳和胚芽中,两产地来源的蓖麻籽中存在显著的丰度差异。且三维重构空间分布中,RCB在连续组织切片中的定位规律较一致。RCB在非靶向数据分析中均归类于贡献度最高的主成分。该MALDI-MSI方法为蓖麻籽溯源研究提供了安全、可靠的新技术途径。