牡丹品种鉴定用ISSR引物的筛选与开发

用于牡丹品种鉴定的DNAISSR-PCR反应体系已经建立。利用DNAISSR分子标记分析少量牡丹品种时,容易获得各品种的特有ISSR标记。然而,中国牡丹品种约有1500个,在小批量品种范围内找到的品种特有ISSR标记有可能出现在其它品种中。因此,利用DNAISSR分子标记对数量庞大的中国牡丹品种进行区分和鉴定时,寻找品种特有标记成为突出的技术难题。标记是由引物通过PCR扩增产生的。因此,关键在于找到理想的ISSR引物。对已知的ISSR引物的筛选未获得良好的PCR扩增结果。报道牡丹鉴定用ISSR引物的设计与开发新途径。     

生长环境与株龄对凤丹当年生果枝的生物量分配的影响

生物量分配研究是了解作物产量形成机制的基础。凤丹是以杨山牡丹（Paeonia ostii）为原种形成的栽培类群,也是新型木本油料作物油用牡丹的主栽品种。该文通过比较凤丹主要产区6龄种群、以及安徽铜陵和上海地区的4、6、8龄种群的当年生果枝大小与生物量分配,探讨环境和株龄对小枝生长与繁殖的影响。结果表明,凤丹植株的果枝数量随株龄增大而上升,不同株龄植株的单个果枝大小和果枝内生物量分配没有明显差异,果枝水平上没有株龄效应,但整株水平上株龄效应明显;果枝大小及生物量各指标之间存在明显相关性,但相关性在种群间变化较大;不同种群的果枝大小及生物量分配差异明显,果枝生物量分配明显随纬度、降水和光辐射强度变化而变化;异速生长方程模拟显示,大部分种群的果枝繁殖与营养生物量或总生物量之间为等速生长关系（斜率=1）,但种群间截距变化较大。这些结果表明栽培环境对凤丹果枝的生长与繁殖有显著效应。     

Global Identification of MicroRNAs and Their Targets in Barley under Salinity Stress

Salinity is a major limiting factor for agricultural production worldwide. A better understanding of the mechanisms of salinity stress response will aid efforts to improve plant salt tolerance. In this study, a combination of small RNA and mRNA degradome sequencing was used to identify salinity responsive-miRNAs and their targets in barley. A total of 152 miRNAs belonging to 126 families were identified, of which 44 were found to be salinity responsive with 30 up-regulated and 25 down-regulated respectively. The majority of the salinity-responsive miRNAs were up-regulated at the 8h time point, while down-regulated at the 3h and 27h time points. The targets of these miRNAs were further detected by degradome sequencing coupled with bioinformatics prediction. Finally, qRT-PCR was used to validate the identified miRNA and their targets. Our study systematically investigated the expression profile of miRNA and their targets in barley during salinity stress phase, which can contribute to understanding how miRNAs respond to salinity stress in barley and other cereal crops.     

牡丹二氢黄酮醇4-还原酶基因PsDFR1的克隆及表达分析

二氢黄酮醇4-还原酶（DFR）是花色素苷合成途径中的关键酶,在植物花色的形成过程中起重要作用。本研究以牡丹品种‘彩绘’为试材,采用RT-PCR和RACE方法从花瓣中获得了一个牡丹DFR基因cDNA全长,命名为PsDFR1,GenBank登录号为HQ283448。序列分析结果表明,PsDFR1全长1242bp,包含一个长为1095bp的开放阅读框,编码364个氨基酸。生物信息学分析显示该基因编码的蛋白具有典型的DFR蛋白功能结构域,包括多个保守的短链脱氢/还原酶家族的特征位点,属于NADB_Rossmann超家族。氨基酸序列比对和系统进化树分析表明,PsDFR1与葡萄、棉花、毛果杨、梨等植物的DFR相似性都在75%以上。实时荧光定量PCR分析表明,PsDFR1在花色素大量积累的花瓣中表达量最高,其次是萼片和雄蕊,再次是叶片,在心皮中表达量最低。     

‘正午’牡丹微繁殖体系的建立

本研究以‘正午’牡丹腋芽为外植体,建立了高效的牡丹微繁殖体系。腋芽的初始培养基为WPM+0.5mg/L BA+0.2mg/L GA3,培养50d后,一个丛生芽平均可切分为13个繁殖体用于增殖培养;增殖培养基为WPM[1668mg/L Ca(NO3)2·4H2O]+0.5mg/L BA+0.2mg/L GA3,以35d为一个继代培养周期,增殖率为3.0,共继代培养7次;生根培养时,先将无根苗在复壮培养基[1/2MS(296mg/L CaCl2)+0.5g/L活性炭]上培养20d,再转入根诱导培养基[1/2 MS(296 mg/L CaCl2)+1.0 mg/L腐胺+1.0 mg/L IBA]培养30d,最后转入根形成培养基[1/2 MS(296mg/L CaCl2)+4.0g/L活性炭]培养20d,其生根率达77.2%;驯化与移栽基质为珍珠岩∶蛭石∶草炭土=1∶1∶1,组培苗移栽成活率高达92.1%。这表明以‘正午’牡丹腋芽建立的微繁殖体系具备规模化商业生产的价值。     

Genome-Wide Identification and Characterization of Germin and Germin-Like Proteins (GLPs) and Their Response Under Powdery Mildew Stress in Wheat (Triticum aestivum L.)

Plant Molecular Biology Reporter - Germin and germin-like proteins (GLPs) play multifaceted roles in plants and participate in signaling processes associated with host–pathogen interactions...     

Identification and Characterization of Novel MicroRNAs from Schistosoma japonicum

Background

Schistosomiasis japonica remains a major public health problem in China. Its pathogen, Schistosoma japonicum has a complex life cycle and a unique repertoire of genes expressed at different life cycle stages. Exploring schistosome gene regulation will yield the best prospects for new drug targets and vaccine candidates. MicroRNAs (miRNAs) are a highly conserved class of noncoding RNA that control many biological processes by sequence-specific inhibition of gene expression. Although a large number of miRNAs have been identified from plants to mammals, it remains no experimental proof whether schistosome exist miRNAs.

Methodology and Results

We have identified novel miRNAs from Schistosoma japonicum by cloning and sequencing a small (18–26 nt) RNA cDNA library from the adult worms. Five novel miRNAs were identified from 227 cloned RNA sequences and verified by Northern blot. Alignments of the miRNAs with corresponding family members indicated that four of them belong to a metazoan miRNA family: let-7, miR-71, bantam and miR-125. The fifth potentially new (non conserved) miRNA appears to belong to a previously undescribed family in the genus Schistosome. The novel miRNAs were designated as sja-let-7, sja-miR-71, sja-bantam, sja-miR-125 and sja-miR-new1, respectively. Expression of sja-let-7, sja-miR-71 and sja-bantam were analyzed in six stages of the life cycle, i.e. egg, miracidium, sporocyst, cercaria, schistosomulum, and adult worm, by a modified stem-loop reverse transcribed polymerase chain reaction (RT-PCR) method developed in our laboratory. The expression patterns of these miRNAs were highly stage-specific. In particular, sja-miR-71 and sja-bantam expression reach their peaks in the cercaria stage and then drop quickly to the nadirs in the schistosomulum stage, following penetration of cercaria into a mammalian host.

Conclusions

Authentic miRNAs were identified for the first time in S. japonicum, including a new schistosome family member. The different expression patterns of the novel miRNAs over the life stages of S. japonicum suggest that they may mediate important roles in Schistosome growth and development.     

Correction to: Genome-wide Identification of Trihelix Genes in Moso Bamboo (Phyllostachys edulis) and Their Expression in Response to Abiotic Stress

Journal of Plant Growth Regulation - The original version of this article unfortunately contained an error in Funding section. The authors would like to correct the error with this erratum.     

Identification and Characterization of ABA-Responsive MicroRNAs in Rice

MicroRNAs(miRNAs) are endogenous non-coding small RNAs that silence genes through mRNA degradation or translational inhibition.The phytohormone abscisic acid(ABA) is essential for plant development and adaptation to abiotic and biotic stresses.In Arabidopsis,miRNAs are implicated in ABA functions.However,ABA-responsive miRNAs have not been systematically studied in rice.Here high throughput sequencing of small RNAs revealed that 107 miRNAs were differentially expressed in the rice ABA deficient mutant,Osabal.Of these,13 were confirmed by stem-loop RT-PCR.Among them,miR1425-5P,miR169 a,miR169n,miR390-5P,miR397 a and miR397 b were up-regulated,but miR162 b reduced in expression in Osabal.The targets of these 13 miRNAs were predicted and validated by gene expression profiling.Interestingly,the expression levels of these miRNAs and their targets were regulated by ABA.Cleavage sites were detected on 7 of the miRNA targets by 5'-Rapid Amplification of cDNA Ends(5'-RACE).Finally,miR162 b and its target OsTREl were shown to affect rice resistance to drought stress,suggesting that miR162 b increases resistance to drought by targeting OsTREl.Our work provides important information for further characterization and functional analysis of ABA-responsive miRNAs in rice.     

“凤丹”牡丹花丝愈伤组织诱导及分化研究

以"凤丹"花丝为外植体，探究了外植体发育时期、基本培养基和植物生长调节剂（plant growth regulators，PGRs）对愈伤组织诱导、增殖和分化的影响，并对不同类型的愈伤组织进行了形态组织学观察。结果表明：花蕾直径为13~19mm时、半透明或白色的幼嫩花丝更适宜愈伤组织的诱导；愈伤组织诱导的最适培养基为SH添加2，4-D 1 mg·L^-1、NAA 2 mg·L^-1及BA 0.2 mg·L^-1，最高诱导率和诱导量分别为99.12%和5.68 g；愈伤组织增殖的最适培养基为SH添加2，4-D 0.5 mg·L^-1、NAA 0.5 mg·L^-1及BA 0.25 mg·L^-1，增殖率为3.86；花丝愈伤组织根据形态结构可分为三类，Ⅰ类愈伤组织具有典型的胚性愈伤组织特征，可进一步分化产生体胚；Ⅱ类愈伤组织和Ⅲ类愈伤组织内部呈现大量维管组织分化，其中Ⅱ类愈伤组织中观察到了不定芽的分化，Ⅲ类愈伤组织未见分化。上述结果表明"凤丹"花丝具有重要再生潜力，是一种良好的新型外植体，对研究建立牡丹体外再生体系具有重要价值。     

The First High-Density Genetic Map Construction in Tree Peony (Paeonia Sect. Moutan) using Genotyping by Specific-Locus Amplified Fragment Sequencing

Genetic linkage maps, permitting the elucidation of genome structure, are one of most powerful genomic tools to accelerate marker-assisted breeding. However, due to a lack of sufficient user-friendly molecular markers, no genetic linkage map has been developed for tree peonies (Paeonia Sect. Moutan), a group of important horticultural plants worldwide. Specific-locus amplified fragment sequencing (SLAF-seq) is a recent molecular marker development technology that enable the large-scale discovery and genotyping of sequence-based marker in genome-wide. In this study, we performed SLAF sequencing of an F1 population, derived from the cross P. ostti ‘FenDanBai’ × P. × suffruticosa ‘HongQiao’, to identify sufficient high-quality markers for the construction of high-density genetic linkage map in tree peonies. After SLAF sequencing, a total of 78 Gb sequencing data and 285,403,225 pair-end reads were generated. We detected 309,198 high-quality SLAFs from these data, of which 85,124 (27.5%) were polymorphic. Subsequently, 3518 of the polymorphic markers, which were successfully encoded in to Mendelian segregation types, and were in conformity with the criteria of high-quality markers, were defined as effective markers and used for genetic linkage mapping. Finally, we constructed an integrated genetic map, which comprised 1189 markers on the five linkage groups, and spanned 920.699 centiMorgans (cM) with an average inter-marker distance of 0.774 cM. There were 1115 ‘SNP-only’ markers, 18 ‘InDel-only’ markers, and 56 ‘SNP&InDel’ markers on the map. Among these markers, 450 (37.85%) showed significant segregation distortion (P < 0.05). In conclusion, this investigation reported the first large-scale marker development and high-density linkage map construction for tree peony. The results of this study will serve as a solid foundation not only for marker-assisted breeding, but also for genome sequence assembly for tree peony.