槐树(Sophora japonica L.)种质资源现状及研究进展

对槐树种质资源在我国的分布和现状,以及山东、山西、河北和河南等地的资源调查收集情况进行了综述;从表型、细胞和生理生化、分子水平3个方面,对槐树种质资源评价的研究进展进行了归纳;以选择育种、人工杂交授粉和现代生物技术育种为例,对种质资源创新利用情况进行了总结,并对工作中存在的问题和发展前景进行了分析。

Selection and Validation of Reference Genes for Normalization of RT-qPCR Analysis in Developing or Abiotic-Stressed Tissues of Loquat (Eriobotrya japonica)

Loquat(Eriobotrya japonica Lindl.)is a subtropical evergreen fruit tree that produces fruits with abundant nutrients and medicinal components.Confirming suitable reference genes for a set of loquat samples before qRT-PCR experiments is essential for the accurate quantification of gene expression.In this study,eight candidate reference genes were selected from our previously published RNA-seq data,and primers for each candidate reference gene were designed and evaluated.The Cq values of the candidate reference genes were calculated by RT-qPCR in 31 different loquat samples,including 12 subgroups of developing or abiotic-stressed tissues.Different combinations of stable reference genes were screened according to a comprehensive rank,which was synthesized from the results of four algorithms,including the geNorm,NormFinder,BestKeeper andΔCt methods.The screened reference genes were verified by normalizing EjLGA1 in each subgroup.The obtained suitable combinations of reference genes for accurate normalization were GAPDH,EF1αand ACT for floral development;GAPDH,UBCE and ACT for fruit setting;EF1α,GAPDH and eIF2B for fruit ripening;ACT,EF1αand UBCE for leaves under heat stress;eIF2B,UBCE and EF1αfor leaves under freezing stress;EF1α,TUA and UBCE for leaves under salt stress;ACT,EF1αand eIF2B for immature pulp under freezing stress;ACT,UBCE and eIF2B for immature seeds under freezing stress;EF1α,eIF2B and UBCE for both immature pulp and seeds under freezing stress;UBCE,TUB and TUA for red-fleshed fruits under cold-storage stress;eIF2B,RPS3 and TUB for white-fleshed fruits under coldstorage stress;and eIF2B,UBCE and RPS3 for both red-and white-fleshed fruits under cold-storage stress.This study obtained different combinations of stable reference genes for accurate normalization in twelve subgroups of developing or abiotic-stressed tissues in loquat.To our knowledge,this is the first report to obtain stable reference genes for normalizing gene expression of abiotic-stressed tissues in E.japonica.The use of the three most stable reference genes could increase the reliability of future quantification experiments.

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+.

Seed Germination Traits of Loquat (Eriobotrya japonica Lindl.) as Affected by Various Pre-Sowing Treatments (Cutting of Cotyledons, Removal of Perisperm, Moist Chilling and/or Exogenous Application of Gibberellin)

The purpose of this study was to investigate the effects of various presowing treatments on the germinability(final germination percentage)and germination rate of loquat seeds in order to increase seedling production in nurseries(applied research)as well as provide answers for important physiological issues related to loquat seeds and their seed coat(basic research).Three experiments were carried out with various pre-sowing treatments.These treatments included full or partial removal of seed coat(perisperm),partial cutting of cotyledons as well as moist chilling at 5℃ for 13 days and/or soaking the seeds in water or 250 ppm gibberellic acid(GA_(3))solution for 24 h.According to the results,cotyledons excision resulted in delayed germination,regardless of the presence or absence of the seed coat in comparison with the decoated seeds that demonstrated the highest germination rate amongst them.In addition,even the partial excision of seed coats affected positively both the germinability and the germination rate,compared to the control-intact seeds.Furthermore,control-intact seeds had a higher germination percentage when exposed to moist chilling independently of the application or not of gibberellin;while the combination of gibberellin application and moist chilling improved both the percentage and the rate of germination of decoated seeds.In conclusion,the role of perisperm(seed coat)in the germination procedure of loquat seeds seems to be important,indicating the existence of seed coat-imposed dormancy on loquat seeds.Finally,the existence of a mild endogenous embryo-dormancy on loquat is also discussed.

Polyphenol Oxidase Inactivation by Microwave Oven and Its Effect on Phenolic Profile of Loquat (<i>Eriobotrya japonica</i>) Fruit

The objective of this research was investigated the effect of polyphenol oxidase microwave treatment on phenolic composition, antioxidant activity and microstructure of loquat fruit. Phenolic profile of methanolic extracts prepared from fresh, and microwave-treated samples were analyzed. Antioxidant activity was also evaluated by 2,2’-azinobis (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS?+) and 1,1-diphenyl-2-picrylhydrazyl (DPPH+) methods. In addition, polyphenol oxidase inactivation was carried out using a response surface methodology to establish the optimal conditions of treatment. The phenolic content of fresh mesocarp was 311 ± 0.60 mg gallic acid equivalents (GAE)/100g dry weight (DW) and that of microwave-treated mesocarp was 1230 ± 0.36 mg GAE/100g DW. Total phenolic content of water/ methanol extract significantly increases after microwave treatment rather than methanolic extract of fresh loquat. Five glycoside phenolics were identified by HPLC-DAD-MS as 3-caffeoylquinic acid, 3-p-coumaroylquinic acid, 5-caffeoylquinic acid and quercetin-3-O-sambubioside. Methanolic extract of microwave-treated mesocarp showed higher antioxidant activity than that of fresh mesocarp. Thus, polyphenol oxidase inactivation by microwave energy preserved the integrity of phenolic compounds as well as antioxidant activity in mesocarp extracts prepared from loquat fruit. It was also noted that phenolics were more abundant in the microwaved samples than in the fresh samples.

枇杷PYL基因家族鉴定及表达分析

脱落酸(ABA)是调节植物生长发育与非生物胁迫的重要激素之一,ABA受体Pyrabactin Resistance 1(PYR)-like(PYL)于信号通路的顶端发挥核心调控作用,但枇杷Eriobotrya japonica的PYL基因家族尚未鉴定。本研究通过生物信息学分析鉴定出枇杷12个PYL基因家族成员,枇杷PYL蛋白分子量为10.47~29.12 kDa,且主要定位于细胞质。枇杷PYL基因家族成员分布在8条染色体上,进化树分析显示PYL基因家族包含三个亚家族,且枇杷EjPYL1与拟南芥AtPYL13亲缘关系较近。枇杷PYL蛋白含有1~6个motif,其中motif3为家族成员共有。枇杷PYL基因家族成员在物种内存在共线性,与拟南芥的物种间共线性有2对。顺式作用元件分析发现枇杷PYL基因启动子含有低温、干旱、光以及激素响应元件。枇杷幼果转录组分析表明,果肉和种子中EjPYL6和EjPYL8基因受低温诱导上调表达。该研究结果将为枇杷ABA信号转导途径的解析和抗冻新品种培育提供理论依据。

耐冬山茶(Camellia japonica L.)花粉活力和柱头可授性研究

采用TTC法测定“长柱耐冬”、“深红皱瓣”、“花团锦簇”、“霜红”4个耐冬山茶品种的花粉活力和寿命，采用联苯胺-过氧化氢法测定4个耐冬山茶品种柱头的可授性。结果表明，‘深红皱瓣’花粉活力在开花期间显著优于‘长柱耐冬’、‘花团锦簇’、‘霜红’3个品种，4个品种的花粉寿命最长可达到37d。4个品种的耐冬山茶柱头在开花1～7d内均具可授性。在1～4d可授性较强，不同的耐冬山茶品种间存在一定差异。

山茶属华东山茶(Camellia japonica L.)研究进展

山茶属华东山茶(Camellia japonica)是我国的传统名花,从生殖生物学、细胞染色体、遗传多样性、育种方法、繁殖技术、化学成分、园林观赏价值、生态价值、食用、油用及药用价值等方面对华东山茶的研究进展进行综述,并对华东山茶未来的育种方向和开发前景进行展望。将现代分子育种与传统育种手段结合运用,同时进一步深入挖掘华东山茶的油用和药用价值并对其合理地开发利用是未来的主要发展方向。

金心黄杨(Euonymus japonica L. cv. aureo-pictus)叶片总蛋白质提取及双向电泳技术

通过比较2种不同的植物叶片总蛋白质提取方法(三氯乙酸/丙酮法和酚法)和优化双向电泳各试验环节,建立金心黄杨叶片总蛋白质的提取方法,以及可以对其蛋白质组进行质谱分析的双向电泳条件。结果采用优化后的酚法进行金心黄杨叶片总蛋白质的提取,IEF蛋白上样量150μg,SDS-PAGE采用12.5%T凝胶,电泳结束后用考马斯亮蓝G-250染色,Melanie 7.0软件分析后得到约274个可分辨蛋白点。

Prospect of the QTL-qSB-9^(Tq) utilized in molecular breeding program of japonica rice against sheath blight

The major QTL-qSB-9^Tq conferring partial resistance to rice （Oryza sativa L.） sheath blight （Rhizoctonia solani Kvhn） has been verified on chromosome 9 of the indica rice cultivar, Teqing. In this study, the prospect of this QTL utilized in molecular breeding program of japonica rice for sheath blight resistance was investigated. Most of the japonica rice cultivars showed lower level of sheath blight resistance than the indica rice cultivars. At the corresponding site of qSB-9^Tq, nine typical japonica rice cultivars from different ecological regions or countries proved to possess the susceptible allele（s）. Introgression of qSB-9^Tq into these cultivars enhanced their resistance level by decreasing sheath blight score of 1.0 （0.5-1.3）, which indicated that qSB-9^Tq had a large potential in strengthening the resistance of japonica rice to sheath blight. The use of the three molecular markers, which were polymorphic between Teqing and many japonica rice cultivars, promotes the application of qSB-9^Tq in a concrete molecular breeding program.

New Diterpenoid Alkaloids from Spiraea japonica var. Ovalifolia

Five new diterpenoid alkaloids. 19-O-deethlyspiramine N (1), deacetylspiramine S (2), spiramidine A (3), spiramidine B (4) and deacetylspiramine F (5) were isolated from the aerial parts of Spirae japonica L. f. var ovalifolia. Their structures were charaterized mainly based on spectral analysis.

Effect of side deep placement of nitrogen on yield and nitrogen use efficiency of single season late japonica rice

Side deep placement of nitrogen plays an important role in improving rice yield and nitrogen use efficiency.Few studies have examined the effects of reducing the times of nitrogen(RTN)application and reducing the nitrogen rate(RNR)of application on rice yield and nitrogen use efficiency under side deep placement of nitrogen in paddy fields.Therefore,a field experiment of RNT and RNR treatments was conducted with nine fertilization modes during the 2018–2019 rice growing seasons in a rice–wheat cropping system of the lower reaches of the Yangtze River,China.Rice yield and nitrogen use efficiency were investigated under side deep placement of nitrogen.We found that under the same nitrogen application rate,the yield of RTN3 increased by 9.64 and 10.18%in rice varieties NJ9108 and NJ5718,respectively,compared with the farmers’fertilizer practices(FFP).The nitrogen accumulation of RTN3 was the highest at heading stage,at 11.30 t ha^(–1)across 2018 and 2019.Under the same nitrogen application rate,the N agronomic use efficiency(NAE),N physiological efficiency(NPE)and N recovery efficiency(NRE)of RTN3 were 8.1–21.28%,8.51–41.76%and 0.28–14.52%higher than those of the other fertilization modes,respectively.RNR led to decreases in SPAD value,leaf area index(LAI),dry matter accumulation,nitrogen accumulation,and nitrogen use efficiency.These results suggest that RTN3 increased rice yield and nitrogen use efficiency under the side deep placement of nitrogen,and RNR1 could achieve the goals of saving cost and increasing resource use efficiency.Two fertilization modes RTN3 and RNR1 both could achieve the dual goals of increasing grain yield and resource use efficiency and thus are worth further application and investigation.

A New Diterpene from Spiraea japonica var. ovalifolia

A new diterpenoid, 15-O-acetylspiraminol (1), was isolated from the aerial parts of Spireae japonica L. f. var. ovalifolia. The structure was charaterized mainly based on spectral analysis.

Effects of chilling tolerance induced by spermidine pretreatment on antioxidative activity,endogenous hormones and ultrastructure of indica-japonica hybrid rice seedlings

Spermidine（Spd） is known to be involved in the regulation of plant responses to chilling stress and counteract the adverse effect of stress conditions.Antioxidant activities,endogenous hormones and ultrastructure change under chilling stress were investigated in indica-japonica hybrid rice seedlings.12-d-old seedlings were subjected to exogenous Spd（1 mmol L^（-1）） and then a chilling stress（6℃,4 d） was induced,followed by a subsequent recovery（25℃,4 d）.Results showed that malondialdehyde（MDA） and proline content were enhanced significantly,whereas shoot fresh and dry weights decreased during chilling stress and after recovery;chlorophyll content of chilling-stressed seedlings increased slightly but declined after recovery;additionally,total soluble sugar,sucrose,fructose and starch contents increased significantly during chilling stress,and only soluble sugar and fructose contents were observed in increase after recovery;chilling stress-induced increases in superoxide dismutase（SOD）,peroxidase（POD） and catalase（CAT） activities,but declined after recovery,and the level of ascorbate peroxidase was lower during chilling stress and after recovery;however,endogenous indole-3-acetic acid（IAA）,zeatin riboside（ZR）,gibberellic acid（GA_3）,and abscisic acid（ABA） levels were induced decreased compared with Spd pretreatment.The microscopic analysis revealed that chilling stress-induced destruction of the chloroplast envelope during chilling stress and increased the number of plastoglobuli along with aberrations in thylakoid membranes after recovery.In contrast,exogenous Spd protected rice seedlings from chilling-induced injuries in terms of lower malondialdehyde,proline and carbohydrates accumulation coupled with increased endogenous hormones metabolism.After recovery,Spd pretreatment chilling-exposed seedlings showed higher activities of antioxidant enzymes and normal physiological function of chloroplasts.These results suggest that Spd could promote effectively chilling tolerance which might be largely attributable to the integrity of cell structure and normal metabolism of endogenous hormones in indica-japonica hybrid rice seedlings.

Primary Research on Protective Effect of Houttuynia cordata Flavonoid and Sophora japonica L. Polysaccharide as Anti-UV Sun-screening Supplement

Houttuynia cordata flavonoid and Sophora japonica L. polysaccharide were investigated to develop anti-UV sun-screening agent through UV wavelength scanning spectrum, antioxidant ability and cell viability after UVB irradiation. The results showed that the extracts had strong UVA and UVB absorption ability and hydroxyl free radical scavenging ability. The S. japonica L. polysaccharide showed strong anti-UVB irradiation ability according to the cell viability experiment. It suggests that the H. cordata flavonoid and S. japonica L. polysaccharide could be combined as a candidate of anti-UV sun-screening supplement and compensate for the shortcomings of chemical sunscreens against UVA ray.

八角金盘水浸提液和GLDA强化马缨丹修复镉污染土壤的效果研究

【目的】探明八角金盘水浸提液(FJ)和谷氨酸N,N-二乙酸四钠(GLDA)复合活化剂强化马缨丹修复镉污染土壤的效果及机理。【方法】采用盆栽试验,研究FJ和GLDA对马缨丹生物量、镉富集特征及根际土壤镉有效性的影响。【结果】GLDA对马缨丹生长无影响,FJ和FJ+GLDA均能促进马缨丹地上部的生长,其地上部干重较CK分别增加了29.26%和19.47%。马缨丹根和茎中的镉含量受FJ的影响较小,而经GLDA和FJ+GLDA处理后分别提高了52.39%、60.66%和34.95%、40.36%。3种活化剂均显著增加了马缨丹地上部的镉富集量,分别较CK增加了53.11%、53.53%和88.79%,其中FJ+GLDA处理的效果最好。土壤DOC含量在3种活化剂处理下均显著提高,分别为CK的1.22、1.34和1.28倍。同时,3种活化剂促进了土壤中可还原态和可氧化态镉向酸提取态镉的转化以及有效态镉含量的增加,其中有效态镉在FJ+GLDA处理下增加了24.26%,优于其他处理。【结论】利用复合活化剂强化马缨丹修复镉污染土壤是可行的。复合活化剂促进了马缨丹的生长,并通过提高土壤DOC含量,增强了土壤镉的有效性,从而促进了马缨丹对镉的吸收和富集,显著提高了马缨丹修复镉污染土壤的效率。

Gametic selection in a doubled haploid population derived from anther culture of indica/japonica hybrid of Oryza Sativa L.

It is one of the key problems for application ofanther culture in hybrid breeding, geneticanalysis, and molecular mapping whether thedoubled haploid (DH) population derived fromanther culture of rice crosses represents a ran-dom array of the microspore population, i.e.whether gametic selection occurs in androgene-sis. A DH population including 132 lines de-

槐提取物和维生素C组合物对紫外照射诱导的HaCaT细胞光损伤的保护作用

本研究旨在探讨槐提取物和维生素C(Vitamin C,V_(C))对紫外线照射引起的HaCaT细胞光损伤的保护作用。通过体外培养HaCaT细胞并将其分为不同组别:对照组(未接受紫外辐照,无槐提取物、V_(C)或二者组合物处理)、紫外线辐射组、125μg/mL槐提取物(接受紫外辐照及槐提取物处理)、125μg/mL维生素C(接受紫外辐照及维生素C处理)以及125μg/mL组合物组(接受紫外辐照及组合物处理,槐提取物和V_(C)质量比为1:1)。采用荧光染色和酶标仪检测技术来评估槐提取物、V_(C)及二者组合物对紫外照射引起的HaCaT细胞产生的活性氧物质(ROS)和线粒体ROS的抑制率。利用免疫荧光染色和酶联免疫吸附法检测槐提取物、V_(C)和组合物对紫外照射引起的HaCaT细胞中环丁烷嘧啶二聚体(Cyclobutane pyrimidine dimers,CPDs)和细胞炎症因子IL-6的表达水平。结果显示,槐提取物、V_(C)和组合物对UVA处理后HaCaT细胞生成的总ROS抑制率分别为38.23%±9.23%、27.20%±6.87%及64.13%±4.08%,与单独使用槐提取物或V_(C)处理相比,组合物显著提高总ROS抑制率(P<0.01)。槐提取物、V_(C)和组合物对线粒体ROS(Mitochondrial DNA,mtROS)的抑制率分别为50.29%±4.92%、38.81%±8.66%及84.74%±3.68%,与单独使用槐提取物或V_(C)处理相比,组合物极显著提高mtROS抑制率(P<0.01)。此外,在UVB照射后,槐提取物、V_(C)及组合物可极显著减少CPDs的生成(P<0.01),这种效应在组合物组效果更为显著(与槐提取物组比较P<0.05;与V_(C)组比较P<0.01)。同时,槐提取物和组合物显著降低UVA照射后炎症因子IL-6的分泌(P<0.01;P<0.05),表明槐提取物及组合物具有抗炎作用。本研究结果表明槐提取物和维生素C组合物能减轻紫外线引起的氧化应激损伤和炎症反应,为未来体内研究和开发具有健康益处的槐提取物和V_(C)组合物提供实验支持。

枇杷鲨烯合酶(Squalene synthase)基因的克隆及序列分析

鲨烯合酶（SQS）是枇杷三萜酸生物合成过程中的关键酶。以枇杷（Eriobotrya iaponicn L．）悬浮培养细胞为材料，采用RT—PCR和RACE方法分离得到枇杷鲨烯合酶Ej-SQS（Squalene svnthase）基因的cDNA全长序列（GenBank，JQ294055），共1775bp，包含了5’非编码区为97bp，开放阅读框为1239bp，3’非编码区为439bp。该cDNA的开放阅读框推定的氨基酸序列（含412个氨基酸）与其它植物SQS具有79％～94％同源性．包含Trans_IPPS_HH保守结构域，可能属于Isoprenoid Biosynthesis enzymes，Class 1家族。为今后利用基因工程调控枇杷细胞悬浮培养物以提取目标产物奠定基础。

NaNO2-Al（NO3）3-NaOH比色法测定枇杷花水提液中总黄酮含量

目的：建立紫外-可见分光光度法测定枇杷花水提液中总黄酮的含量,为枇杷花新食品原料的申报奠定基础。方法：以芦丁为对照品,采用NaNO2-Al（NO3）3-NaOH显色反应体系,对显色剂的浓度与用量进行考察,筛选出最佳显色条件,在505 nm处用紫外-可见分光光度法测定样品中总黄酮的含量。结果：采用该方法可以准确地测定枇杷花水提液中总黄酮的含量,总黄酮在16.4898.88μg/mL范围内线性关系良好,回归方程为y=6.470 2x-0.003 7,R2=0.999 4,样品平均回收率为95.56%,RSD为3.1%。结论：该方法简便、快捷、准确,可作为枇杷花水提液总黄酮含量测定的方法。