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Accurate quantification of 3'-terminal 2'-O-methylated small rnas by utilizing oxidative deep sequencing and stem-loop RT-qPCR

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Frontiers of Medicine 2022年第2期16卷 240-250页

作者： Yan Kong Huanhuan Hu Yangyang Shan Zhen Zhou Ke Zen Yulu Sun Rong Yang Zheng Fu Xi Chen Nanjing Drum Tower Hospital Center of Molecular Diagnostic and Therapy State Key Laboratory of Pharmaceutical BiotechnologyJiangsu Engineering Research Center for MicroRNA Biology and BiotechnologyNJU Advanced Institute of Life Sciences(NAILS)Institute of Artificial Intelligence BiomedicineSchool of Life SciencesNanjing UniversityNanjing 210023China Department of Urology Drum Tower HospitalMedical School of Nanjing UniversityInstitute of UrologyNanjing UniversityNanjing 210008China Department of General Surgery Drum Tower HospitalMedical school of Nanjing UniversityNanjing 210008China Research Unit of Extracellular RNA Chinese Academy of Medical SciencesNanjing 210023China

The continuing discoveries of novel classes of RNA modifications in various organisms have raised the need for improving sensitive,convenient,and reliable methods for quantifying RNA *** particular,a subset of small rnas,including micrornas(mirnas)and Piwi-interacting rnas(pirnas),are modified at their 3'-terminal nucleotides via 2'-***,quantifying the levels of these small rnas is difficult because 2'-0-methylation at the RNA 3'-terminus inhibits the activity of polyadenylate polymerase and T4 RNA *** two enzymes are indispensable for RNA labeling or ligation in conventional miRNA quantification *** this study,we profiled 3'-terminal 2'-0-methyl plant mirnas in the livers of rice-fed mice by oxidative deep sequencing and detected increasing amounts of plant mirnas with prolonged oxidation *** further compared the efficiency of stem-loop and poly(A)-tailed RT-qPCR in quantifying plant mirnas in animal tissues and identified stem-loop RT-qPCR as the only suitable ***,stem-loop RT-qPCR was superior to poly(A)-tailed RT-qPCR in quantifying 3'-terminal 2'-0-methyl pirnas in human seminal *** summary,this study established a standard procedure for quantifying the levels of 3'-terminal 2'-0-methyl mirnas in plants and *** measurement of the 3'-terminal 2'-0-methylation of small rnas has profound implications for understanding their pathophysiologic roles in biological systems.

关键词： small rnas 2'-0-methylation sequencing RT-qPCR

Species-specific regulatory pathways of small rnas play sophisticated roles in flower development in Dimocarpus longan Lour.

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Horticultural Plant Journal 2023年第2期9卷 237-249页

作者： Bo Liu Guanliang Li Chengjie Chen Zaohai Zeng Jing Xu Jisen Zhang Rui Xia Yuanlong Liu State Key Laboratory for Conservation and Utilization of Subtropical Agro-Bioresources South China Agricultural UniversityGuangzhouGuangdong 510640China Guangdong Laboratory for Lingnan Modern Agriculture South China Agricultural UniversityGuangzhouGuangdong 510640China Key Laboratory of Biology and Germplasm Enhancement of Horticultural Crops in South China Ministry of Agriculture and Rural AffairsSouth China Agricultural UniversityGuangzhouGuangdong 510640China Center for Genomics and Biotechnology Haixia Institute of Science and TechnologyCollege of Life SciencesFujian Agriculture and Forestry UniversityFuzhouFujian 350002China

Flower development plays vital role in horticultural ***-transcriptional regulation via small rnas is important for plant flower *** uncover post-transcriptional regulatory networks during the flower development in Dimocarpus longan Lour.‘Shixia’,an economically important fruit crop in subtropical regions,we collected and analyzed sRNA deep-sequencing datasets and degradome libraries Apart from identifying mirnas and phased siRNA generating loci(PHAS loci),120 hairpin loci,producing abundant srnas,were identified by in-house *** results suggested that 56 miRNA-target pairs,2221-nt-PHAS loci,and 111 hairpin loci are involved in posttranscriptional gene silencing during longan reproductive ***-specific or species-specific post-transcriptional regulatory modules have been unveiled,including miR482-PHAS and ***482-PHAS might be involved in longan flower development beyond their conserved roles in plant defense,and miRN15 is a novel miRNA likely associated with a hairpin locus(HPL-056)to regulate strigolactone receptor gene DWARF14(D14)and the biogenesis of phasirnas from *** small rnas are enriched in flower buds,suggesting they are likely involved in post-transcriptional regulatory networks essential for longan flower development via the strigolactone signaling pathway.

关键词： Dimocarpus longan Lour small rnas Flower development Post-transcriptional regulation

Roles of small rnas in crop disease resistance

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Stress Biology 2021年第1期1卷 65-79页

作者： Jun Tang Xueting Gu Junzhong Liu Zuhua He National Key Laboratory of Plant Molecular Genetics CAS Center for Excellence in Molecular Plant SciencesInstitute of Plant Physiology&EcologyChinese Academy of SciencesShanghai 200032China University of the Chinese Academy of Sciences Beijing 100049China State Key Laboratory of Conservation and Utilization of Bio-Resources in Yunnan and Center for Life Sciences School of Life SciencesYunnan UniversityKunmingChina.

small rnas(srnas)are a class of short,non-coding regulatory rnas that have emerged as critical components of defense regulatory networks across plant *** sRNA-based technologies,such as host-induced gene silencing(HIGS),spray-induced gene silencing(SIGS),virus-induced gene silencing(VIGS),artificial microRNA(amiRNA)and synthetic trans-acting siRNA(syn-tasiRNA)-mediated RNA interference(RNAi),have been developed as disease control strategies in both monocot and dicot plants,particularly in *** review aims to highlight our current understanding of the roles of srnas including mirnas,heterochromatic sirnas(hc-sirnas),phased,secondary sirnas(phasirnas)and natural antisense sirnas(nat-sirnas)in disease resistance,and srnas-mediated trade-offs between defense and growth in *** particular,we focus on the diverse functions of srnas in defense responses to bacterial and fungal pathogens,oomycete and virus in ***,we highlight the application of sRNA-based technologies in protecting crops from *** research perspectives are proposed to develop new srnas-based efficient strategies to breed non-genetically modified(GMO),diseasetolerant crops for sustainable agriculture.

关键词： small rnas HIGS Crop diseases RNAi RNAi-based technology

番茄果实中成熟衰老相关small rnas的最新研究进展

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农业生物技术学报 2016年第2期24卷 270-279页

作者：左进华王清鞠正曹东艳朱本忠罗云波高丽朴国家蔬菜工程技术研究中心果蔬农产品保鲜与加工北京市重点实验室中国农业大学食品科学与营养工程学院

番茄(Lycopersicon esculentum)是研究果实成熟衰老的模式材料,其成熟是一个多基因调控的高度协调的遗传调控过程,伴随着色泽、质地、风味、香气和其他代谢等许多显著的生理生化变化。近年来,small rnas作为一种新型的转录后调控因子受... 详细信息

番茄(Lycopersicon esculentum)是研究果实成熟衰老的模式材料,其成熟是一个多基因调控的高度协调的遗传调控过程,伴随着色泽、质地、风味、香气和其他代谢等许多显著的生理生化变化。近年来,small rnas作为一种新型的转录后调控因子受到越来越多的关注,几乎参与了动植物中所有的生理和生化过程。在植物中参与生长发育、果实成熟、生物和非生物胁迫反应等过程。本文系统地综述了番茄果实中small rnas的分离鉴定现状,对番茄果实中与乙烯信号途径、颜色、风味、软化等代谢途径相关的small rnas进行了系统总结,并展望了未来small rnas在果实成熟过程中的相关研究方向。

关键词：番茄果实成熟衰老 small rnas 研究进展

micrornas和高表达small rnas在水稻两个亚种及正反交F1代中的杂种优势分析

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microRNAs和高表达small RNAs在水稻两个亚种及正反交F1代中的杂...

作者：陈芳芳北京协和医学院

学位级别：硕士

杂种优势是杂交F1代在生物量、株高、生长发育速度、繁殖力等农艺性状方面优于其父母本的现象。虽然杂种优势已经被研究和运用超过了一个世纪,但是它的分子机制仍然不甚明了。我们的研究着重于了解small RNA在杂种优势中发挥的作用。通... 详细信息

杂种优势是杂交F1代在生物量、株高、生长发育速度、繁殖力等农艺性状方面优于其父母本的现象。虽然杂种优势已经被研究和运用超过了一个世纪,但是它的分子机制仍然不甚明了。我们的研究着重于了解small RNA在杂种优势中发挥的作用。通过一个专门检测small rnas表达水平的芯片体系,检测了水稻两个亚种日本晴(Nipponbare)和93-11以及它们正反交F1代中所有已知的micrornas和高表达的那部分small rnas的表达水平。在所有检测的1141条small rnas中,正反交中分别有140和157条small rnas有明显差异表达,差异表达的比例在12-13%。加性效应和非加性效应的四种情况(高显性、低显性、超高显性、超低显性)均有发现。正反交F1代均以非加性效应为主。且相对于父母本的中亲值,表达下调的small rnas数目明显多于表达上调的。我们也发现有15个mirnas的表达水平在正反交中相对于中亲值表现了出了相反的趋势。生物信息学手段预测显示水稻micrornas的targets参与了各种生物学途径,且表达有差异和无差异的micrornas,其targets并未表现有明显区别。我们的研究表明small rnas在水稻的杂种优势表现中起了重要作用,为进一步探索和发现杂种优势的分子机制提供了理论依据。

关键词：杂种优势 small rnas 差异表达加性效应非加性效应

Prediction of candidate small non-coding rnas inAgrobacterium by computational analysis

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The Journal of Biomedical Research 2010年第1期24卷 33-42页

作者： Tingting Zhao Ren Zhang Mingbo Wang The Laboratory Center for Basic Medical Sciences Nanjing Medical University Nanjing 210029 Jiangsu Province China Department of Btologtcal Sciences University of Wollongong Wollongong NSW 2522 Australia CSIRO Plant Industry Canberra ACT2602 Australia

small non-coding rnas with important regulatory roles are not confined to eukaryotes. Recent work has uncovered a growing number of bacterial small rnas （srnas）, some of which have been shown to regulate critical cellular processes. Computational approaches, in combination with molecular experiments, have played an important role in the identification of these srnas. At present, there is no information on the presence of small non-coding rnas and their genes in the Agrobacterium tumefaciens genome. To identify potential srnas in this important bacterium, deep sequencing of the short RNA populations isolated from Agrobacterium tumefaciens C58 was carried out. From a data set of more than 10,000 short sequences, 16 candidate srnas have been tentatively identified based on computational analysis. All of these candidates can form stem-loop structures by RNA folding predictions and the majority of the secondary structures are rich in GC base-pairs：：Some are followed by a short stretch of U residues, indicative of a rho-independent transcription terminator, whereas some of the short rnas are found in the stem region of the hairpin, indicative of eukaryotic-like srnas. Experimental strategies will need to be used to verify these candidates. The study of an expanded list of candidate srnas in Agrobacterium will allow a more complete understanding of the range of roles played by regulatory rnas in prokaryotes.

关键词： small non-coding rnas small rnas Agrobacterium turnefaciens genome solexa sequencing technology

Regulatory rnas in Bacillus subtilis:A review on regulatory mechanism and applications in synthetic biology

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Synthetic and Systems Biotechnology 2024年第2期9卷 223-233页

作者： Anqi Peng Guobin Yin Wenjie Zuo Luyao Zhang Guocheng Du Jian Chen Yang Wang Zhen Kang The Science Center for Future Foods Jiangnan UniversityWuxi214122China The Key Laboratory of Carbohydrate Chemistry and Biotechnology Ministry of EducationSchool of BiotechnologyJiangnan UniversityWuxi214122China The Key Laboratory of Industrial Biotechnology Ministry of EducationSchool of BiotechnologyJiangnan UniversityWuxi214122China

Bacteria exhibit a rich repertoire of RNA molecules that intricately regulate gene expression at multiple hierarchical levels,including small rnas(srnas),riboswitches,and antisense ***,the majority of these regulatory rnas lack or have limited protein-coding capacity but play pivotal roles in orchestrating gene expression by modulating transcription,post-transcription or translation *** and redesigning these regulatory RNA elements have emerged as pivotal strategies in the domains of metabolic engineering and synthetic *** previous investigations predominantly focused on delineating the roles of regulatory RNA in Gram-negative bacterial models such as Escherichia coli and Salmonella enterica,this review aims to summarize the mechanisms and functionalities of endogenous regulatory rnas inherent to typical Gram-positive bacteria,notably Bacillus ***,we explore the engineering and practical applications of these regulatory RNA elements in the arena of synthetic biology,employing *** as a foundational chassis.

关键词： Regulatory RNA devices small rnas Riboswitches Synthetic biology Bacillus subtilis

RNA silencing:From discovery and elucidation to application and perspectives

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Journal of Integrative Plant Biology 2022年第2期64卷 476-498页

作者： Jian-Hua Zhao Hui-Shan Guo State Key Laboratory of Plant Genomics Institute of Microbiologythe Chinese Academy of SciencesBeijing100101 China CAS Center for Excellence in Biotic Interactions University of the Chinese Academy of SciencesBeijing100049 China

RNA silencing(or RNA interference,RNAi)is a conserved mechanism for regulating gene expression in *** discovery of natural trans-kingdom RNAi indicated that small rnas act as signaling molecules and enable communication between organisms in different *** phenomenon and potential mechanisms of trans-kingdom RNAi are among the most exciting research *** better understand trans-kingdom RNAi,we review the history of the discovery and elucidation of RNAi *** on canonical RNAi mechanisms,we summarize the major points of divergence around RNAi pathways in the main eukaryotes’kingdoms,including plants,animals,and *** review the representative incidents associated with the mechanisms and applications of trans-kingdom RNAi in crop protection,and discuss the critical factors that should be considered to develop successful trans-kingdom RNAi-based crop protection strategies.

关键词： crop protection gene expression RNA silencing small rnas trans-kingdom RNAi

GmDFB1,an ARM-repeat superfamily protein,regulates floral organ identity through repressing siRNA-and miRNA-mediated gene silencing in soybean

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Journal of Integrative Plant Biology 2024年第8期66卷 1620-1638页

作者： Jie Li Wenxiao Zhang Qing Lu Jiaqi Sun Chuang Cheng Shiyu Huang Shuo Li Qiang Li Wei Zhang Chuanen Zhou Bin Liu Fengning Xiang The Key Laboratory of Plant Development and Environmental Adaptation Biology Ministry of EducationSchool of Life SciencesShandong UniversityQingdao 266237China State Key Laboratory of Crop Gene Resources and Breeding Institute of Crop SciencesChinese Academy of Agricultural SciencesBeijing 100081China

The development of flowers in soybean(Glycine max)is essential for determining the yield potential of the *** silencing pathways are involved in modulating flower development,but their full elucidation is still ***,we conducted a forward genetic screen and identified an abnormal flower mutant,deformed floral bud1-1(Gmdfb1-1),in *** mapped and identified the causal gene,which encodes a member of the armadillo(ARM)-repeat *** small RNA sequencing(sRNA-seq),we found an abnormal accumulation of small interfering rnas(si rnas)and microRNA(mirnas)in the Gmdfb1 *** further demonstrated that GmDFB1 interacts with the RNA exosome cofactor SUPER KILLER7(Gm SKI7).Additionally,GmDFB1 interacts with the PIWI domain of ARGONAUTE 1(GmAGO1)to inhibit the cleavage efficiency on the target genes of s *** enhanced gene silencing mediated by siRNA and miRNA in the Gmdfb1 mutants leads to the downregulation of their target genes associated with flower *** study revealed the crucial role of GmDFB1 in regulating floral organ identity in soybean probably by participating in two distinct gene silencing pathways.

关键词： ARM-repeat superfamily protein ARGONAUTE 1 SUPER KILLER 7 flower development gene silencing small rnas soybean