Combining Transcriptomics and Metabolomics to Uncover the Effects of High-Energy Lithium-Ion Beam Irradiation on Capsicum annuum L.

Hot pepper(Capsicum annuum L.)is consumed as one of the oldest domesticated crops all over the world.Although mutation breeding using radiation has been performed in hot peppers,little is known about the comparative analysis of mutagenic effects at the molecular level by ion beam irradiation.To comprehend the response mechanism of hot pepper to the ion beam,we used a mutant with favorable economic characteristics induced by lithium-ion beam irradiation to investigate the biological effects.The results indicated that the lithium-ion beam had a positive effect on important agronomic traits,particularly yield unit,but had a negligible effect on the photosynthetic rate of hot pepper,with a specific influence on chlorophyll b rather than chlorophyll a.By RNA-Seq analysis,671 up-regulated and 376 down-regulated genes were identified as differentially expressed genes(DEGs)between irradiated and unirradiated hot pepper.Based on GO and KEGG network analysis,the auxin metabolic process was the common pathway in these two networks.A total of 118 potential reactive oxygen species(ROS)scavenging genes and 262 signal transduction genes were identified,suggesting a balance between antioxidant enzymes and enhanced ROS transduction.The amounts of 15 metabolite,involved in GABA pathways,secondary metabolism,carbohydrate metabolism,shikimate pathways,TCA cycles,nitrogen metabolism,glycerol metabolism and acetate pathways,were significantly changed in the ion beam irradiated sample.These results highlighted that the enriched pathways could play important roles in response to ion beam irradiation in hot pepper plants.In summary,these data provide valuable information for future research on ion beam irradiation and genomic studies in hot pepper.

Effect of Different Culture Conditions on Sporulation Quantity of Phytophthora capsici in Capsicum annuum L. var. dactylus M. in Xunhua

[Objective] The paper was to explore the effect of different culture conditions on sporulation quantity of Phytophthora capsici in Capsicum annuum L.var.dactylus M.in Xunhua.[Method] The effects of light hours,pH,medium and temperature on sporulation quantity during isolation and culture process of P.capsici were studied.[Result] The sporulation quantity of P.capsici under the conditions of 24 h/day light,pH 7.0,potato medium（PSA） and 30 ℃ was the largest,and pH,basal medium and temperature had greater impact on sporulation quantity.[Conclusion]The study laid foundation for the study on natural incidence condition of C.annuum in Xunhua.

五彩椒(Capsicum annuum L.)的组织培养研究

以抗寒性较强的五彩樱桃椒为试验材料,对观赏椒的组织培养进行了研究。试验表明,最佳外植体为茎尖,茎尖的最佳消毒时间为3min;最佳诱导培养基为MS+6-BA5mg·L-1+NAA1mg·L-1;最佳分化培养基为MS+6-BA5mg·L-1+NAA0.1mg·L-1,最佳生根培养基为1/2MS+IBA0.25mg·L-1。

Construction of Mutant Population and Analysis of Dwarf Mutants in "6421"(Capsicum annuum L.)through EMS Mutagenesis

[Objective] The aim was to improve the genetic property of peppers, the mutant population of Capsicum annuum L cultivar ＂6421＂ was constructed. [Method] The seeds of ＂6421＂ were treated with 0.2% to 1.2% ethyl methane sulfonate to identified LD50, and then 10 000 LD^o of treated seeds were sowed to construct mutant population. The agronomic characters and genetic regularity of dwarf mutants in M4 generation were analyzed. [Result] Our results showed that GR and SSR were 45.2% and 40.2% respectively at 1.0% EMS, close to LD50, with GI （17.6） and seed Ⅵ （19.7） being half of that of control; 562 M4 mutants were identified in 2015, and the mutation could be characterized according 11 major categories and 32 subcategories; Simultaneously, we found that plant height, plant width, diameter of mainstem, length of main-stem, the number of main-stem nodes and branch of lines E29, E58, E142 and E312 were all significantly different from that of the control. The mutation of lines E29, E58 and E312 was all controlled by a single recessive gene. [Conclusionl The study first created a pepper mutant population, which provides not only the germplasm resources for further breeding but also direct and effective materials for genomic study of the pepper.

Cloning and characterization of CaGID1s and CaGAI in Capsicum annuum L.

Fruit set and development are affected by many phytohormones, including gibberellin. Little is known regarding molecular mechanism underlying gibberellin mediated fruit set and development especially in Capsicum. Three gibberellin recep- tors, CaGIDlb.1, CaGIDlb.2 and CaGIDlc, and a DELLA protein, CaGAI, have been identified in Capsicum annuum L. During the fruit development, the expression level of CaGIDlc was low, and the expression fold change is mild. However, CaGIDlb. 1 and CaGIDlb.2 were relatively higher and more acute, which indicates that CaGIDlb. 1 and CaGID1b.2 may play an important role in fruit pericarp, placenta and seed. Ectopic expressions of CaGIDlb. 1, CaGIDlb.2 and CaGIDlc in Arabidopsis double mutant gidla gidlc increased plant height, among which CaGIDlb.2 had the most significant effect; CaGAI reduced plant height in double mutant rga-24/gai-t6, having a similar function to AtGID1 and AtGAI in stem elon- gation. Yeast two-hybrid （Y2H） and bimolecular fluorescence complementation （BiFC） assays indicated that CaGIDlb.1 and CaGID1 b.2 interact with CaGAI in a GA-dependent manner, while CaGIDlc interacts with CaGAI in a GA-independent manner. Our study reveals the key elements during gibberellin signaling in Capsicum and supports the critical importance of gibberellin for Capsicum fruit set and development.

Combining Ability Analyses of Net Photosynthesis Rate in Pepper (Capsicum annuum L.)

In perspective of breeding high-yield hybrid pepper varieties, combining ability analysis of net photosynthesis rate at different phases of flowering and fruit setting in pepper was made with 15 cross combinations from 6 parents by （1/2） n （n- 1） diallel crosses. There are relatively large differences not only in general combining ability （GCA） effect among different parents and at different phases of flowering and fruit setting, but also in specific combining ability （SCA） effect among different hybrids. There are relatively large GCA effects in late parents but relatively less GCA effects in early parents. No obvious laws have been found in the relationship between SCA effects and maturity of hybrids. Variances of SCA are larger than those of GCA. Heritability is less but influence of environment is larger. Correlation analysis of combining ability between net photosynthesis rate and agronomic character or resistances to main diseases has showed that correlation coefficients of GCA are relatively large at the medium phase and the late phase of flowering and fruit setting. Net photosynthesis rate is more relative to leaf characters and fruit characters. Correlation coefficients of SCA are relatively large at the early phase and the late phase of flowering and fruit setting. Net photosynthesis rate is more relative to leaf characters and plant characters at the early phase but to plant characters and fruit characters at the late phase. Correlation coefficients of SCA between net photosynthesis rate and resistances to main diseases are larger than those of GCA. The combining abilities of net photosynthesis rate at different phases of flowering and fruit setting are positively correlated with those of yield per plant. The combining ability is an important parameter of breeding of high photosynthesis hybrid pepper varieties.

Effect of continuous negative pressure water supply on the growth, development and physiological mechanism of Capsicum annuum L.

Effects of continuous negative pressure water supply on water consumption, growth and development, as well as physiological mechanism and quality of Capsicum annuum L. were investigated in this paper. Meanwhile, the optimal negative pressure water supply conditions for growth of C. annuum L. were screened out to achieve the goals of water conservation, high yield and high quality, thus providing theoretical foundation for its field production. The pot experiment within the greenhouse was utilized; the continuous negative pressure water supply was adopted; the four treatments, artificial watering（CK）, –5 k Pa（T1）, –10 k Pa（T2）, and –15 k Pa（T3） were set; and the daily water consumption, yield, as well as the biomass, nitrate reductase, root activity, vitamin C, capsaicin, and nutrient uptakes of nitrogen（N）, phosphorus（P） and potassium（K） during various stages of its growth were determined. Compared with CK, when the water supply pressure was controlled at –5 to –15 k Pa in the experiment, the total water consumption of C. annuum L. reduced by 53.42 to 67.75%, the total water consumption intensity reduced by 54.29 to 67.14%, and the water use efficiency increased by 12.66 to 124.67%. The N accumulation in a single strain of C. annuum L. from the color turning stage to the red ripe stage increased by 15.99 to 100.55%, respectively, compared with that of CK; the P accumulation increased by 20.47 to 154.00% relative to that of CK, and the K accumulation increased by 64.92 to 144.9% compared with that of CK. Compared with CK, C. annuum L. yield was remarkably improved by 13.79% at T1, and contents of vitamin C, capsaicin as well as carotenoids at all growth stages were enhanced by 13.42–147.01%, 11.54–71.01%, and 41.1–568.06%, respectively. Nitrate reductase activity, root activity and chlorophyll（a＋b） were markedly increased by 335.78–500%, 79.6–140.68% and 114.95–676.19%, respectively, from immature stage to full ripe stage. Adopting the continuous negative pressure water supply for C. annuum L. has a significant water-saving effect, and the water supply pressure being stable at –5 k Pa contributes to its growth and development, improves yield, enhances root activity, promotes nutrient uptake, and improves its quality, thus achieving the effects of water conservation, high yield, high quality and high efficiency.

Purification and Biochemical Characterization of a Protease Inhibitor Ⅱ Family from Jalapeno Pepper(Capsicum annuum L.)

Capsicum annuum L. was initially domesticated in Mexico and northern Central America, and represented an ancient Neotropical plant food complex. The purpose of this paper is to report the isolation and purification of a novo-member of a protease inhibitor from jalape&ntildeo pepper (Capsicum annuum L.) (PIJP). The molecular weight of PIJP inhibitor is 5.95 kDa with 56 amino acids and 6 Cys residues with high inhibitory activity to trypsin with a Ki value of 95 nM. This inhibitor according to the alignment with homologous from NCBI and Pfam databases is a member of proteinase inhibitors II. It is worthwhile to mention a major compositional difference between the proteinase inhibitor II families which have 8 Cys residues. PIJP is the first purified proteinase inhibitor, member of this family with only 6 Cys residues.

Processing of Snack of Goat Chub Chili(Capsicum annuum L.)with Formulations of Tomato and Tomato-Marjoram

The cultivation of goat chili has potentially increased in Chile, due to different varieties of gourmet products made with it;for this reason, new industrial projects are sought to the growing volume to adapt them to the new trends in consumption habits, mainly focused on achieving an attractive meal, a fast preparation and consumption, while nutrients are provided to offer some benefits for human health. The “snacks” products that include the use of fruit satisfactorily respond to these requirements, in addition to providing marketing alternatives to the surplus of fruit exports. For this reason, the present test, consisting of the processing of snacks based on goat chub chili with the addition of tomato and marjoram, was carried out to increase the palatability and the sensorial quality of the final product. The following formulations were used: only chili (T1);chili and tomato (T2);chili, tomato and marjoram (T3). The latter one was characterized by a good flavor and aroma. A large amount of the fruit water content had been removed by drying it with forced hot air. A product without chemical preserves was obtained, with physical and chemical parameters similar to those required by similar foods existing on the market today. In sensorial terms, the formulation to which marjoram was added showed good sensorial and acceptability attributes, highlighting them from the rest of the treatments. The snack color was maintained after the dehydration process, according to panelists.

气孔在辣椒(Capsicum annuum L.)子叶和下胚轴上的动态变化

以辣椒幼苗(Capsicum annuum L.)为材料,研究了辣椒生长发育过程中子叶的上、下表皮和下胚轴上、下端1 cm处的气孔密度及表皮细胞密度的动态变化。结果表明:(1)辣椒的气孔由肾形保卫细胞组成,无副卫细胞;子叶的表皮细胞呈无规则形,下胚轴的表皮细胞呈长方形或椭圆形;(2)辣椒子叶上、下表皮的气孔密度和表皮细胞密度分别在辣椒种子出芽后第6天和第3天达到最大,之后随子叶的生长逐渐减小,下表皮的气孔密度和表皮细胞密度明显大于上表皮的;(3)辣椒下胚轴上、下端1 cm处的气孔密度随下胚轴的生长变化不明显,但是表皮细胞密度随下胚轴的生长逐渐减小,下胚轴上端的气孔密度和表皮细胞密度明显的大于下胚轴下端的。

基于加权关联网络分析的辣椒(Capsicum annuum L.)果实颜色转变相关基因的鉴定

辣椒属于茄科(Solanaceae)辣椒属(Capsicum L.)一年生或多年生植物,其播种面积和产值在中国均居蔬菜首位。果色是辣椒众多经济性状中最直观的性状之一,直接影响人们的购买选择。辣椒果色受多种因素调控,相关色素生物合成的分子调控还需进一步解析。本研究以果实发育过程中呈现5种颜色的辣椒种质HNCA0076为实验材料,进行了生理检测、转录组测序以及WGCNA分析。在果实5个不同颜色时期,叶绿素含量变化较大,类胡萝卜素含量整体呈现递增趋势,类黄酮含量无明显变化,花色苷含量在紫色果期后逐渐降低,橙色果期后又逐渐升高。RNA测序数据质量满足进一步分析要求。4个转色期的上调(下调)表达基因数分别为826(276)、390(588)、1248(1800)和1528(2485),随着果实的逐步发育,相邻比较组合中的差异基因的数量逐渐增加。差异表达基因经GO富集分析,主要集中在分子功能中的四吡咯结合、血红素结合、转移酶活性、氧化还原酶活性、铁离子结合和水解酶活性等过程;KEGG富集分析主要富集在类胡萝卜素生物合成、类黄酮生物合成和淀粉、蔗糖代谢等信号通路,其中类胡萝卜素代谢途径的3个基因在紫色时期向黄色时期的转变过程中表达上调,类黄酮代谢途径的11个基因在黄色时期向红色时期的转变过程中表达下调。4个对比组合差异基因分别注释到8877、8736、8689、8573个转录因子,共同注释到8049个转录因子,包括AP2、F-box、MYB和UDPGT等类型。利用非冗余的差异表达基因进行了加权关联网络分析,关联到4个与叶绿素、类胡萝卜素、花色苷、类黄酮含量相关的模块,进一步分析表明p450(LOC107859314)、Pkinase(LOC107839192、LOC107875415)、F-box(LOC107863894)和zf-MYND(LOC107852593)等29个转录因子可能参与辣椒果实转色的调控。本研究为进一步探索辣椒果实色素形成过程中的关键调控因子以及分析相关色素生物合成的调控提供了重要数据。

Diallel Crossing Analyses of Resistance to Main Diseases in Pepper(Capsicum annuum L.)

Fifteen capsicum combinations were made with 6 parents by （1/2）n（n-1） diallel crossing. Genetic parameters in the resistance to TMV, CMV, phytophthora blight, bacterial spot of these combinations were studied by Hayman. The results indicated that the resistance to TMV, CMV and bacterial spot conformed genetically to the “additive-dominant” model but the resistance to phytophthora blight did not and significant epistatic dominance effect existed in it. F1 hybrid＇s resistance to CMV was controlled by homozygous dominant gene （s）, but resistance to bacterial spot by heterozygous one （s）. There were little, or no sum of dominant effect and genomes controlling the dominant expression of F1 hybrids in its phytophthora blight resistance.

AFLP Analysis of Mitochondrial DNA from a Cytoplasmic Male Sterility Line and its Maintainer in Pepper (Capsicum annuum L.)

Cytoplasmic male sterility (CMS) is a maternally inherited trait characterized by the inability of a plant to produce functional pollen, which is widespread among higher plants. CMS system is a valuable tool for plant

辣椒(Capsicum annuum L.)果实发育及品质相关新microRNA的鉴定与分析

MicroRNA(miRNA)是一种非编码的小RNA,在各种生物过程中起着重要的调节作用。前人研究表明,模式植物中miRNA与果实发育相关。然而,人们对辣椒(Capsicum annuum L.)果实品质及发育相关的miRNA了解尚不清楚。本研究利用二代测序技术(Next-generation sequencing technology)比较了不同品种辣椒果实在不同发育时期的小RNA。利用miRDeep2软件从4个样本中共鉴定出了59个已知miRNA和310个新miRNA。预测了新miRNA的靶基因,共656个,并对其中402个靶基因进行了注释。GO分析和KEGG途径表明,某些靶基因与淀粉糖、氨基糖以及核糖代谢相关。qPCR表明一些miRNA的表达模式与其靶向基因相反,该结果为今后研究miRNA调控果实发育和品质形成提供了重要基础。

辣椒(Capsicum annuum L.)耐涝突变体相关功能基因的表达分析

EMS诱变辣椒亲本‘S15’获得1个耐涝突变体‘RW15’。前期研究认为,保护酶和内源激素代谢相关的8个基因与其耐涝特性相关。本研究通过对不同水涝处理时间的‘S15’和‘RW15’的根和叶进行基因实时定量分析,以探讨其在耐涝机制中的作用。研究发现,水涝胁迫下,生长素诱导蛋白相关的Cap.ARATH,乙烯响应相关的Cap.RAP2、MYB家族相关的Cap.MYB1R1和过氧化物酶Cap.POD相关的4个基因表达量等7个基因表达量在突变体中显著升高,另1个与过氧化物酶Cap.POD相关Capana02g003649基因表达量却下降;另外,同一处理时间的‘RW15’对水涝胁迫响应时间快及响应程度高于‘S15’,Cap.MYB1R1表达量最高,且胁迫前后表达差异达到6.94倍。研究表明,突变体‘RW15’主要通过提高内源激素和保护酶代谢等相关功能基因的表达达到耐涝能力的增强,该研究对突变体耐涝机制解析提供数据了参考。

几丁质和flg22诱导的辣椒幼苗先天免疫生理响应特征

【目的】为明确几丁质和鞭毛蛋白衍生肽flg22诱导的辣椒幼苗先天免疫生理响应特征,探讨辣椒先天免疫生理响应与辣椒抗多种病害的关系。【方法】以5个四川本地辣椒品种幼苗为试材,鉴定它们对青枯病和疫病病情指数和抗性水平,采用水培法培育幼苗并进行外源几丁质和flg22处理,检测各品种不同诱导时间下幼苗根系生长、气孔孔径、胼胝质沉积、活性氧(ROS)积累和SOD、CAT活性,以及先天免疫相关基因表达量的变化,综合评价其生理响应及其与抗病性关系。【结果】(1)各品种幼苗青枯病和疫病病情指数以‘川腾10号’最低,抗病性最强,以‘本地条椒’最高,抗病性最弱。(2)外源几丁质和flg22抑制了各品种幼苗根系生长速率,诱导离体叶片气孔闭合,促进叶片细胞壁胼胝质沉积加厚,ROS含量持续增加,SOD和CAT活性不断提高。各品种幼苗先天免疫生理响应指标的平均隶属函数值以‘川腾10号’最高,‘本地条椒’最低,并且平均隶属函数值与疫病、青枯病病情指数均具有显著负相关性。(3)外源flg22和几丁质诱导‘川腾10号’幼苗先天免疫相关基因CaWRKY22、CaMAPK7和ChiIV3显著上调表达。【结论】外源flg22和几丁质可诱导辣椒幼苗先天免疫生理响应,且响应程度强弱在品种间存在差异,依据生理响应指标通过隶属函数可综合评价辣椒品种的抗病水平;‘川腾10号’的平均隶属函数值最高,多抗性水平最好,这与其先天免疫相关基因CaWRKY22、CaMAPK7和ChiIV3的显著上调表达有关。

辣椒EIL基因家族的鉴定及其在盐碱胁迫下的表达分析

EIL(Ethylene-insensitive 3-like)基因在参与植物乙烯信号通路的转导和生长发育过程中发挥着重要作用。为了解辣椒EIL基因家族成员信息,利用生物信息学手段对辣椒EIL基因家族成员的理化性质、蛋白质结构、系统进化、基因结构、保守域、启动子顺式作用元件及CaEILs基因在不同组织部位和不同非生物胁迫条件下的表达模式进行分析。同时使用实时荧光定量PCR(qRT-PCR)对辣椒叶片中CaEILs在盐碱胁迫下的表达模式进行研究。结果表明,辣椒9个CaEILs分布在6条染色体上,氨基酸数目、蛋白质理论分子质量和脂肪系数分别介于209~677、23.77~76.07 ku和63.10~87.75,且主要为酸性、亲水的不稳定性核蛋白。系统进化分析显示,CaEILs被分成了4个亚家组,9个CaEILs在根、茎、叶、花蕾、花中具有不同程度的表达,而低温、高温、高盐、干旱胁迫下均能诱导CaEILs的表达,其对以上非生物胁迫均具有不同程度的响应。此外,利用qRT-PCR对盐碱胁迫下辣椒叶片中CaEILs进行了检测,结果发现,随着处理时间的延长,CaEIL1、CaEIL2、CaEIL4、CaEIL5、CaEIL8的表达量整体呈上升趋势,而CaEIL3、CaEIL6、CaEIL7、CaEIL9的表达量整体呈下降趋势。

Residue Dynamics of Mancozeb in Capsicum annuum Linn and Soil

[ Objective ] The paper was to study the residue dynamics of mancozeb in Capsicum annuum Linn and soil. [ Method ] Field spraying test and indoor analysis were carried out in three places of Changsha, Guangzhou and Beijing City during 2008 -2009 to study the digestion dynamics and the final residue of man- cozeb in C. annuum and soil, and gas chromatography was used for quantitative analysis. [ Result]The average recovery rates of mancozeb in C. annuum and soil were 86.59%-93.03% and 92.57%-95.14%, respectively. Mancozeb had rapid digestion in C. annuum and soft with the half-life of 2.29 -4.65 and 4.01 - 4.65 d, respectively. When metalaxyl-M,mancozeb water dispersible granule was sprayed in C. annuum according to the recommend dose （ spraying for no more than three times with the spraying interval of 5 d）, the final residue of maneozeb in C. annuum collected in the third day after the last spraying was less than 2 mg/kg.

Responses of the Anthocyanin and Osmolyte Contents of the Capsicum annuum Cultivars Planted in Wenshan Prefecture of Yunnan Province to the Drought Stress Simulated by PEG-6000

[Objective] The aim of this study was to compare the drought resistance difference of the main Capsicum annuum cultivars planted in Wenshan Prefecture of Yunnan Province. [Method] The total anthocyanin, soluble sugar, soluble protein and free proline contents of the leaves of the five main C. annuum cultivars planted in Wenshan Prefecture, i.e., ZSZ75-1, ZSZ49-1-1, 12WS-18-1, 12ZH01 and ZS130, under the drought stress simulated by PEG-6000 were studied by using spectrophotometry, and the total osmoregulation abilities of the leaves were evaluated by using subordinate function. [Result] Under the drought stress simulated by PEG-6000, the total anthocyanin contents of the veins, mesophylls and whole leaves of the five cultivars were all as： ZSZ75-1〉ZSZ49-1-1〉12WS-18-1〉12ZH01〉ZS130, the soluble sugar contents as： 12ZH01 〉12WS-18-1 〉ZS130〉ZSZ49-1-1 〉ZSZ75-1, the soluble protein contents as： ZSZ75-1〉12ZH01〉ZSZ49-1-1〉12WS-18-1〉ZS130, and the free proline contents as： ZS130〉ZSZ75-1 〉12WS-18-1 〉ZSZ49-1-1 〉12ZH01. Furthermore, the differences among the total anthocyanin contents of the veins, mesophylls and whole leaves of the five cultivars all reached the extremely significant levels, whereas the differences among the contents of the soluble sugar, soluble protein and free proline did not reach the significant levels, the correlation degrees among the contents of the total anthocyanin, soluble sugar, soluble protein and free proline of different cultivars were also inconsistent, and the total osmoregulation abilities of the five cultivar leaves were as： ZSZ75-1 〉7SZ49-1-1 〉12WS-18-1 〉ZS130〉12ZH01. [Conclusion] The anthocyanin accumulation in the specific organs of the five C. annuum cultivars in Wenshan Prefecture is beneficial to the strengthening of the leaf osmoregulation abilities under drought stress, favoring the drought tolerance of the cultivars.

PAP3 Regulates Stamen but Not Petal Development in Capsicum annuum L.

Pepper flowers are hermaphroditic; the plant's male sterility trait is characterized by its inability to produce pollen grains. In the ABC model of flower development, B-function genes play roles in petal and stamen development in the angiosperm. In this study, a B-class gene designated as PAP3 (Gen Bank accession no. HM104635) was isolated in pepper. The gene encoded 226 amino acids and shared high similarity with the MADS-box protein family, with a conservative MADS domain and semiconservative K domain. Furthermore, the expression of PAP3 was abundant only in petals and anthers but not in leaves. A functional study employing virus-induced gene silencing(VIGS) showed that knockdown of PAP3 led to the shriveling of pollen grains and male sterility; however, it did not affect petal development. These results suggest an essential role for PAP3 in the development of the pepper stamen and in contributing to the variation of floral traits.