Identification, mRNA Expression, and Functional Analysis of Chitin Synthase 1 Gene and Its
关于差异表达基因的几种筛选方法
【关键词】 mRNA 差异显示；基因表达；DNA 微阵列 【摘要】 多种因素导致的基因差异性表达与疾病的发生发展密切有关，分离 差异表达基因，对于研究细胞生命过程的调节机制及致病机制具有重要意义. 20 世纪 90 年代以来，先后出现了 mRNA 差异显示 PCR（mRNA DDRTPCR）、代表性差异 分析（RDA）、抑制性消减杂交（SSH）、基因表达连续性分析（SAGE）和 DNA 微阵 列（DNA microarray）等多种分析差别表达基因的方法. 我们对以上方法的原理、 基本步骤及其应用进行简要综述. 【关键词】 mRNA 差异显示；基因表达；DNA 微阵列 0 引言 随着各类基因组计划的相继完成，人类面临的更艰巨的任务是研究基因功能活 动，也就是说基因组序列分析仅仅代表了遗传信息复杂性的一个层次，而遗传信息 有序地、时相地表达则是决定生物体及其行为的另一个层次. 所以，发现不同生物 体及其组织在各种状态下（正常状态、发育、衰老、损伤及疾病）差异表达的基因 具有十分重要的意义，于是差异表达基因筛选技术应运而生. 目前，基因表达差异 的分析通常用稳定状态下 mRNA 的丰度高低及有无进行比较. 差异表达基因有两个 含义，即表达基因的种类变化和基因表达量的变化. 传统的基因分析方法如 Northern 杂交、斑点杂交等存在费时、费力的缺点，已不适宜进行大规模基因表 达分析研究的需要. 因此随着分子生物学的发展，出现了大量新方法，按其技术特 点可分为三类：①以杂交为基础的技术，包括 Northern blotting， Slmapping/Rnase 保护、抑制性消减杂交和 DNA 微阵列；②以 PCR 为基础的技术， 如差异显示 PCR（DDPCR）、代表性差异分析（RDA）；③以测序为基础的技术，如 表达序列标签（EST）、基因表达连续性分析（SAGE）等. 我们对目前主要的差异 表达基因的筛选方法作一综述. 1mRNA 差异显示 PCR(differential display PCR，DDPCR) mRNA 差异显示 PCR 又称为差别显示反转录 PCR（differential display reverse transcription PCR， DDRTPCR）. DDRTPCR 技术［1-3］最早于 1992 年 出现，可以用于分离在不同的真核细胞中差异表达的 cDNA 并加以克隆. 其原理是 将两种细胞的 mRNA 逆转录后进行 PCR 扩增. PCR 3′端引物序列是针对 mRNA 的 poly(A)尾设计的，一般是 11 个 T 再加上两个碱基，这样 12 种 3′端引物 (T11AA，T11AC， T11AG，T11AT，T11CA，T11GA，T11CC，T11CG，T11CT，T11GC， T11GG，T T11GT)就可以与所有 mRNA 的 poly(A)尾匹配；5′端引物是随机引物， 一般为 10 个碱基，因此产生一些不同长度的 cDNA 片段，电泳后比较两者的差别而 得到差异表达基因的 cDNA. 但这个方法存在许多严重的缺陷，它的 5′端随机引物 一般常有 2～3 个碱基不能与 cDNA 模板完全匹配，而且 PCR 反应中随机性、偶然性 比较大，容易形成非特异性扩增而造成高的假阳性率，这就使下游的筛选工作很巨 大. 理论上此方法可以检测到 95％以上的转录体，但由于引物序列的随机性和竞 争性模板结合位点的存在，很难确定实际的原始 RNA 丰度. 尽管有上述缺陷，但由 于其实验步骤较简单，此方法在实际工作中应用仍较多，例如用于筛选在肿瘤发
有参转录组测序 英文
Materials and methodsRNA extraction,library construction,and RNA-SeqRNA was extracted from the samples a ccording to the instruction manual of the TRIzol reagent(Invitrogen,Carlsbad,CA)RNA concentration and purity was mesured using the NanoDrop2000Spectrophotometer(Thermo Fisher Scientific,Wilmington, DE).RNA integrity was assessed using the RNA Nano6000Assay Kit of the Agilent Bioanalyzer2100system System(Agilent Technologies,CA,USA).High-quality RNA were sent to Biomarker Technologies Corporation(Beijing,China)for cDNA libraries construction and sequencing.mRNA were purified by the interaction of the poly(A)tails and magnetic oligo(dT)beads.RNA sequencing libraries were generated using the NEBNext®Ultra RNA Library Prep Kit for Illumina(New England Biolabs,Ipswich,MA,U.S.A.)with multiplexing primers,according to the manufacturer’s protocol.The cDNA library was constructed with average inserts of200 bp(150~250bp),with non-stranded library preparation.The cDNA was purified using a AMPure XP Beads(Beckman Coulter,Inc.).The short cDNA fragments were subjected to end repair,adapter ligation.Then,the suitable fragments were selected by Agencourt AMPure XP beads(Beckman Coulter,Inc.),and enriched by PCR amplification. Sequencing was performed via a paired-end125cycle rapid run on the Illumina HiSeq2500.Transcriptome analysis using reference genome-based reads mappingLow quality reads,such as only adaptor,unknown nucleotides>5%,or Q20<20% (percentage of sequences with sequencing error rates<1%),were removed by perl script. The clean reads that were filtered from the raw reads were mapped to mice(XXXXX拉丁名)genome(参考基因组版本)using Tophat2(Kim,Pertea et al.2013)software.Gene expression levels were estimated using FPKM values(fragments per kilobase of exon per million fragments mapped)by the Cufflinks software(Trapnell,Williams et al.2010) Identification of differential gene expressionDESeq(Anders and Huber2010)and Q-value were employed and used to evaluate differential gene expression between queen and worker honeybee.After that,geneabundance differences between those samples were calculated based on the ratio of the FPKM values.The false discovery rate(FDR)control method was used to identify the threshold of the P-value in multiple tests in order to compute the significance of the differences.Here,only gene with an absolute value of log2ratio≥2and FDR significance score<0.05were used for subsequent analysis.Sequence AnnotationGenes were compared against various protein database by BLASTX,including the National Center for Biotechnology Information(NCBI)non-redundant protein(Nr) database,Swiss-Prot database with a cut-off E-value of10-5.Genes were retrieved based on the best BLAST hit(highest score)along with their protein functional annotation.To annotate the gene with gene ontology(GO)terms,the Nr BLAST results were imported into the Blast2GO program(Conesa,Gotz et al.2005).GO annotations for the genes were obtained by Blast2GO.This analysis mapped all of the annotated genes to GO terms in the database and counted the number of genes associated with each term. Perl script was then used to plot GO functional classification for the unigenes with a GO term hit to view the distribution of gene functions.The obtained annotation was enriched and refined using TopGo(R package).The gene sequences were also aligned to the Clusters of Orthologous Group(COG)database to predict and classify functions(Tatusov, Galperin et al.2000).KEGG pathways were assigned to the assembled sequences by perl script.。
219387083_牛DSCAM_基因的基因组印记和DNA_甲基化状态分析
第46卷第3期2023年5月河北农业大学学报JOURNAL OF HEBEI AGRICULTURAL UNIVERSITYVol.46 No.3May.2023牛DSCAM基因的基因组印记和DNA甲基化状态分析靳兰杰1，霍浩楠1，张银蛟1，李冬杰2，陈玮娜3，张 萃1，李世杰1（1. 河北农业大学 生命科学学院, 河北 保定 071000；2.河北科技大学 食品与生物学院, 河北石家庄 050018；3.河北大学 中医学院，河北 保定 071000）摘要：为鉴定牛DSCAM基因的印记状态以及DNA甲基化修饰在调控印记表达中的作用，本研究以牛胎盘和成年牛组织（心、肝、脾、肺、肾、肌肉、脂肪和大脑）为试验材料，利用基于单核苷酸多态（SNP）的RT-PCR直接测序法分析DSCAM基因的印记状态，采用亚硫酸氢盐测序法分析基因启动子区的甲基化状态。
结果发现，在DSCAM基因的第32个外显子上存在1个A/G杂合的SNP位点（rs136908595），利用该SNP位点区分亲本等位基因发现，在被检测的成年牛组织中，DSCAM基因为双等位基因表达；而在胎盘中DSCAM基因为母源等位基因表达。
对牛DSCAM基因启动子区及第一个外显子处402 bp的CpG岛甲基化进行分析，发现在单等位基因表达的胎盘中，2条亲本链间存在差异甲基化区，而在双等位基因表达的组织中，未发现差异甲基化区。
结果表明，DNA甲基化修饰参与调控牛DSCAM基因的胎盘特异性父源印记，可为深入探讨牛DSCAM基因功能和调控机制提供参考依据。
关 键 词：DSCAM基因；基因组印记；等位基因表达；牛；差异甲基化区中图分类号：S823；S813.3 开放科学（资源服务）标识码（OSID）：文献标志码：AAnalysis of DNA methylation on DSCAM gene in different bovine tissues JIN Lanjie1, HUO Haonan1, ZHANG Yinjiao1, LI Dongjie2, Chen Weina3, ZHANG Cui1, LI Shijie1（1.College of Life Science, Hebei Agricultural University, Baoding 071001, China; 2.College of Food Science andBiology, Hebei University of Science and Technology, Shijiazhuang 050018, China; 3.College of Medical Science,Hebei University, Baoding 071001, China ）Abstract: The DNA methylation status of DSCAM gene in cattle were analyzed to determine the role of DNAmethylation on gene expression. The allelic expression of DSCAM gene was analyzed in bovine placenta and somatictissues including heart, liver, spleen, lung, kindy, muscle, fat and brain by a SNP-based RT-PCR products directsequencing method. The DNA methylation status of CpG island in promoter region of DSCAM gene was analyzedby bisulfite sequencing. An A/G SNP (rs136908595) was found on the exon 32 of bovine DSCAM gene and usedto distinguish the parental alleles. The results showed that DSCAM gene was biallelically expressed in all detectedbovine somatic tissues, including heart, liver, spleen, lung, kidney, muscle, fat and brain. In bovine placenta, DSCAM收稿日期：2023-01-06基金项目： 国家自然科学基金(31372312)；河北省自然科学基金(C2020204004，C2022201058)；河北省人社厅引进留学人员资助项目(C20200332).第一作者：靳兰杰（1997—），女，河北保定人，硕士研究生，主要从事动物基因组学与表观遗传学研究.E-mail:**********************通信作者：李世杰（1971—），女，河北保定人，教授，主要从事动物分子遗传与表观遗传学研究. E-mail:*********************本刊网址：文章编号：1000-1573(2023)03-0091-06DOI：10.13320/ki.jauh.2023.004792第46卷河北农业大学学报gene was maternally expressed based on the SNP (rs136908595). The DNA methylation was analyzed on the CpG island (402 bp) in the promoter region and the first exon of DSCAM gene. A differentially methylated region (DMR) was detected in placenta. However, DMR was not found in spleen, kidney and brain tissues. The results indicated that DNA methylation played a role in the placenta-specific paternal imprinting of bovine DSCAM gene, which provides a reference for further study of the function and regulation mechanism of bovine DSCAM gene ,which provides a reference for further study of the function and regulation mechanism of bovine DSCAM gene and regulation mechanism of bovine DSCAM gene.Keywords: DSCAM gene; genomic imprinting; allelic expression; cattle; DMR1 材料与方法1.1 供试样品的采集印记状态分析包括成年荷斯坦奶牛的组织和胎盘。
生物信息学考试参考题目
1. 在NCBI进行BLAST序列比对时，需要输入查询序列的信息，以下错误的格式是（ C ）A. 序列的accession numberB. 序列的giC. 序列对应基因的IDD. FASTA 格式的序列2. 下面这段序列是: ( B )>gi||ref|| Drosophila melanogaster RNA-binding protein 4 CG9654-RA, transcript variant A (Rbp4),mRNAGGATTTTCTTGCCTGTCA TTCAA TTTGTGGTTGGCTTCACCTGAGTGCTGTAGT。
A. DNA序列B. RNA序列C. 蛋白质序列D. 基因3. ExPASy上的工具软件ProtParam提供的是哪一种类型的服务？（ B ）A．蛋白质三级结构分析B．蛋白质序列理化性质预测C．蛋白质二级结构分析D．跨膜结构分析4. 假设你有两条远相关的蛋白，为了比较它们，最好利用下列哪个记分矩阵（A ）A. BLOSUM45或PAM250B. BLOSUM45或PAM1C. BLOSUM80或PAM250D. BLOSUM10或PAM15. 构建系统发生树，应利用CA. BLASTB. FASTAC. UPGMAD. Entrez6. 下面这段蛋白质序列是什么格式? ( D )>gi|4506183|ref|| proteasome alpha 3 [Homo sapiens]MSSIGTGYDLSASTFSPDGRVFQVEYAMKA VENSSTAIGIRCKDGVVFGVEKLVLS KL YEEGSNKRLFNVDRHVGMA V AGLLADARSLADIAREEASNFRSNFGYNIPLKHLADRV AMYVHAYTL YSA VRPFGCSFMLGS。
A. GBFFB. TEXTC. PDBD. FASTA7. 直系同源物概念为（A ）A．不同物种中具有一路先人的同源序列B．具有较小的氨基酸一致性可是有较大的结构相似性的同源序列C．同一物种中由基因复制产生的同源序列D．同一物种中具有相似的而且一般是冗余功能的同源序列8. 美国NIH保护提供的DNA序列数据库是：（ A ）A. GenBankB. ProteinC. dbESTD. dbSNP9. 高分派对片段的英文缩写为（A ）A. HSPB. HMPC. HCPD. HDP10. BLAST比对结果报告中有一统计数值E值，该值大小与匹配度的关系是（ B ）A. 值越小说明匹配度越低B. 值越小说明匹配度越高C. 二者无内在关系D. 以上说法都不对11. NCBI提供了大量的序列分析工具，其顶用来寻觅DNA序列潜在的蛋白质编码区的工具是：（A ）A. ORF FinderB. BLASTC. Scan PrositeD. SAGEmap12. Entrez是哪个网站数据库的检索系统（A ）A．NCBIB．PROSITEC．EBID．PDB13. 若是想找一个和查询蛋白远源的蛋白质，下面哪一种方式最可能成功？ BA．采用PHI-BLAST，因为你能自己选择一个和搜索蛋白质有关的信号序列B．采用PSI-BLAST，因为那个算法利用位点特异性打分矩阵最为敏感C．采用BLASTP，因为你能够调整你的打分矩阵从而使得搜索敏感度最大D．采用专门的物种数据库，因为他们中可能含有这种远源序列。
分子生物学名词解释（Molecularbiologicalterms）
分子生物学名词解释（Molecular biological terms）The beanmail polar tropical fish is set to exitDouban-douban-douban-douban-douban-douban-douban-douban-dou ban-douban-douban-douban stationDouban searchThe home page of my douban my group of the city browse discoveryExplanation of molecular biology terms.Confused の detectiveThe 2010-05-21 20:43:54 from: confused の detective (if you want to let me live please give me happy pain)Title: molecular biology noun explanation (personal arrangement, only a lot of ~ exams are risky, review should be cautious ~ true love life, far away from the point of birth)Meristematic name solutionProbe: molecular hybridization and the tagged nucleotide chain with specific sequences of nucleotide nucleotide nucleotides can be used to detect specific genes in nucleic acid samples.Molecular hybridization: a technique for qualitative or quantitative analysis of DNA or RNA using the basic properties of DNA degeneration and renature.Gene chip: the support of a specific piece of DNA that is closely aligned in a unit area.Gene library: a clone group that contains the entire DNA sequence of an object in a lifetime.CDNA library: it is contain a tissue cells under certain conditions all mRNA expression by the reverse transcription and synthesis of cDNA sequence of clone population, it is stored in the form of cDNA fragments with the tissue cell gene expression information.Genomic DNA library: a clone population stored in the form of fragments of DNA (including all coding and non-coding regions) of the genome of a living organism.Transgenic technology: gene transfer technology is used to integrate the target genes into the fertilized egg cells or embryonic stem cells, then the cells are imported into the animal's uterus to develop into individual technology.Transgenic: the gene that is being imported in transgenic technologyTransgenic animals: the receptor animals that are genetically engineered to be genetically engineeredNuclear transfer technique: an individual cell nucleus of an animal is introduced into the activated egg cell of another individual to develop into an individual, namely, clone.Gene elimination: a technique for removing certain genes in animals based on homologous recombination.Functional cloning: cloning the pathogenic gene by understanding the function of a pathogenic gene.Location cloning: gradually narrowing the range from the chromosomal localization of a pathogenic gene and finally cloning the gene.Gene diagnosis: direct detection of gene structure and its expression level is normal, so as to diagnose the disease.Gene therapy: an exogenous gene that functions as a defective cell can be used to correct or compensate for its genetic defects to achieve therapeutic purposes.Viral oncogene: a type of gene that is present in tumor viruses (mostly retroviruses) that can cause malignant transformation of the target cell.Proto-oncogene: is the oncogene in normal cells, and its expression products regulate the normal growth and differentiation of cells. When activated, can cause cell growth differentiation abnormality, form tumour. Also called cell carcinoma genes.Oncogene: a normal gene in a living organism or in a cell, which controls the growth and differentiation of cells. Cell carcinogenesis can only be caused when its structure changes or expresses an abnormality.Tumor suppressor gene: inhibits the proliferation and proliferation of cells and thus inhibits the genes of tumor formation.Transformation action: by automatically obtaining or artificially supplying exogenous DNA, the cells obtain a new genetic phenotype, which is called transformation.Conjugation: when a cell or bacterium interacts with the bacteria, the plasmid DNA can be transferred from one cell (bacteria) to another (bacteria). This DNA transfer is called the conjugation.Transduction function: when the release of the virus from infected cells, infected again another cell, occurred in donor cells and DNA transfer and recombination between receptor cells is called transduction.Plasmid: small ring double stranded DNA molecule outside the bacterial chromosomeHomologous recombination: a recombination between the homologous sequences, which is also called fundamental recombinationSite specific recombination: the integration of integrase catalysis between the specific sites of two DNA sequences.Transposition: the translocation or rearrangement of genes mediated by insertion sequence and transposon is referred toas transpositionTransposon: a discrete sequence of repeated sequences that can be transferred from one chromosome site to another.Clone: a collection of identical copies or copies from the same ancestor.Cloning: the process of obtaining the same copy, i.e. asexual reproduction.DNA cloning: the method of application enzymology to reorganize the target gene and carrier DNA in vitro, transform or transfect host cells, and gain a large number of genes. It is also called gene cloning and recombinant DNAGenetic engineering: the methods and related work for gene cloning are described as genetic engineering.Compatibility terminal: some of the restriction enzyme recognition sequences are not exactly the same, but after cutting DNA, they produce the same sticky end, which is called the compatibility terminal.Restriction endonuclease: a specific sequence of DNA that identifies a DNA, and an enzyme that cuts double strand DNA around the identification site or its surroundings.CDNA: transcriptional synthesis of single stranded DNA that complements mRNA. Double stranded cDNA can be synthesized with single stranded cDNA as template and polymerized.Genomic DNA: a complete set of genetic information (chromosomes and mitochondria) of a cell or organism.Gene carrier: some of the DNA molecules used to reproduce or express a meaningful protein for the purpose of carrying a target gene.Gene: a genetic base unit located on a chromosome that carries a DNA fragment of a specific genetic information that can encode a single biological product, including RNA and polypeptide chains.Genome: a complete set of genetic information from a living organism, which is the entire genetic information or whole gene that a cell or virus carries.Gene expression: the process of transcription and translation of genesTime specificity: the expression of a specific gene takes place in a certain chronological order according to the function, which is called the time specificity of gene expressionSpace specificity: in the whole process of individual growth, a certain gene product appears in the order of individual tissue space, which is called the spatial specificity of gene expressionHousekeeping genes: some genes continue to be expressed in almost all cells of an individual, often referred to ashousekeeping genesConstitutive gene expression: usually a gene expression similar to the expression of a butler gene, also known as basic expression.Coordinate expression: under certain mechanism control,A set of genes related to function, no matter how they are expressed, should be coordinated and expressed in a coordinated manner.Trans action: the protein factor expressed by one gene interacts with the specific cis-acting component of another gene to regulate its expression. This conditioning is known as the trans action.Cis: protein factor can recognize, regulate the sequence of its own genes, regulate the expression of its own genes, and call it cis.Self - control: regulation of protein is generally used in automrna, inhibiting the synthesis of itself, and self-controlMonocistron: a coding gene transcribed to generate an mRNA molecule that translates into a polypeptide chain.Protein biosynthesis is the process of synthesizing proteins in the sequence of nucleotides in mRNA molecules.S - D sequence: in prokaryotes initiation codon upstream 8 to13 AUG nucleotide site there are 4-9 consensus sequence, high in purine bases, is with small ribosome binding sites of mRNA, known as the S - D sequence.Ribosomal circulation: the peptide chain is extended continuously in the nucleoprotein body, also known as prolongation. This includes carry, peptide and peptide.Polyribosome: the polymer that mRNA forms with multiple nucleosomes is called polyribosomeMolecular partner: the molecular partner is a nonnatural conformation that identifies a type of protein in the cell that can recognize the right folding of the functional domain and the whole protein.Cistron: a genetic unit that codes for a polypeptide.Signal sequence: all sorting signals exist in the targeted delivery of protein structure, mainly was the specific N terminal amino acid sequence, may guide protein metastasize to the appropriate target cells, the sequences are called signal sequence.Open reading framework: the sequence of nucleotide sequences from the mRNA initiation codon AUG to the termination codon.The degeneracy of the genetic code: an amino acid can have two or more codons coded for it, a feature known as the degeneracy of the genetic code.Enhancers: a sequence of DNA that binds specific gene regulation proteins to promote the expression of specific genes near or far away. The distance of the enhancement subscriptional start point varies greatly, but it always ACTS on the most recent promoter.Transcription is the process by which an organism USES DNA as a template to synthesize RNAStructural genes: segments of RNA that are transcribed from DNA molecules called structural genesAsymmetric transcription: in the genome, the genes are transcribed only by the genes of different developmental timing, conditions and physiological needs of the cell. In the DNA molecule double strand, a strand is used as a template for transcription, and the other strand is not transcribed. The template chain is not always on the same chain.Manipulation: transcription is not continuous. Each transcriptional block can be considered as a transcriptional unit, called the operator. The manipulators include several structural genes and their upstream regulatory sequences.Cis-acting elements: the sequence of DNA that is involved in transcriptional regulation at the beginning of the transcription starting point, consisting of promoters, enhancers, and silences.Anti-type action factor: the ability to recognize and combine the homeopathic components, and reverse the transcriptionaleukaryotic proteins that are transcribed by other genes.Transcription factor: in the trans action factor,The direct or indirect combination of RNA polymerase is called transcription factor.Exon: a sequence of nucleic acid sequences of mature rnas in eukaryotic organisms that appear in the fault gene and its primary transcription products.Intron: linear expression of the partition gene in eukaryotes and the sequence of nucleic acids removed during the splicing process.Nuclease: RNA that has enzymatic activity is called a nucleaseReproduction: refers to the generation of genetic material, the process of synthesizing subchain DNA by the mother chain DNA.Semi - reserved replication: when DNA biosynthesis, the mother chain DNA is unwound into two single strands, each acting as a template by the base pairing rule, and the subchain complemented by the template. The DNA of the daughter cell, a single strand is fully accepted from the parent, and the other single strand is completely resynthesized. The DNA of the two subcells is identical to the parental DNA base sequence. This replication method is called semi-retained replication.Bidirectional replication: when the prokaryote replicates, the DNA dissolves the chain from the starting point to the twodirections, forming the opposite of the two directions of the replication fork, called bidirectional replication.Initiator: a compound structure formed at the beginning of DNA replication, containing helicases, DnaC proteins, primers, and DNA replication initiation regions.Replicator: the unit that completes the replication independently, from the replication point to the replication endpoint.Okazaki fragment: a discontinuous fragment formed in the following chain replication during DNA replication.The double strand of DNA is divided into two segments, each acting as a template, and the subchain lengthens the formed y-font structure along the template to be called a replication fork.Half-discontinuous replication: the lead chain replicates continuously and the attendant chain discontinuous replication is called semi-discontinuous replication.Reverse transcription: the process of synthesizing DNA with RNA as a template for reverse transcriptase.Telomere: the structure of the end of the linear DNA molecule of the eukaryotic biosomatic chromosome, which makes the ends of chromosomes become granule.Transcriptional initiation complex: a compound formed by thebinding of the prokaryotic RNA polymerase, the transcriptional pppGpN - product and template DNA.Boundary sequence: the eukaryotic introns start with GU at 5 'end and AG is 3' end. 5 '- GU... Ag-oh3 is called a boundary sequence, also known as a splice interface.Splicing: removing introns from RNA molecules so that exons can be connected together.Promoter: a sequence of DNA in the upstream of the transcription starting point of RNA polymerase. If the RNA polymerase is combined with it, it can initiate transcription.Central rule: the law of transmission of genetic information from DNA to RNA to protein.Transcriptase: transcriptional complex, which is composed of the nuclease of RNA polymerase and the product of its template DNA, transcribed.Silencing: the negative regulating element in the eukaryotic element, which ACTS as a repressor when it binds the specific protein factor.Gene: the structure of eukaryotic gene, by a number of coding and non-coding area interval with each other, but embedded in a row, after connected to again unless the coding regions, can translate the continuous complete protein amino acid composition. These genes are called broken genes.[figure]DavidDavid (I'm looking for the book direction)It is also the common touch of biochemistryYour response? Who? Who? Who? Who? Who? , ah ha... Good thing ~ plus go> medical student's home groupLatest topic:Molecular biology noun explanation (individual, only a lot more to test a risk... (confused の detective)N reasons you attended medical school (breba)I was depressed. (hh)How microbes learn?Everybody work first or take the exam first. (MaoMaoQ)My man is studying medicine (summer)The division room chooses the direction, especially lost... (ssu xiaawn)Are there any graduate students in Concorde? Seek counsel (summercool)> to report bad information2005-2010 , all rights reserved about , contact us · disclaimer · help center · douban service (API) · mobile phone douban · brand club。
胰腺癌诊断和预后关键生物标志物的筛选鉴定和综合分析
doi:10.3971/j.issn.1000-8578.2020.19.0743·临床研究·胰腺癌诊断和预后关键生物标志物的筛选鉴定和综合分析柳兴源1，李菁媛2，杨静1Identification and Integrated Analysis of Key Biomarkers for Diagnosis and Prognosis of Pancreatic AdenocarcinomaLIU Xingyuan 1, LI Jingyuan 2, YANG Jing 11. Basic Medical College, Jinzhou Medical University, Jinzhou 121000, China;2. Department of Pharmacy, The First Af filiated Hospital of Jinzhou Medical University, Jinzhou 121000, ChinaAbstract: Objective To screen and identify key genes and pathways involved in the progression of pancreatic adenocarcinoma (PAAD) and to perform an integrated analysis. Methods GEO2R was commonly used to screen and identify differentially expressed genes (DEGs) in GSE15471, GSE16515, GSE28735 and GSE62165, and the GO and KEGG pathway analyses were performed with DA VID database. Then a protein-protein interaction (PPI) network was constructed using a STRING database, and Cytoscape and GEPIA were used to identify the Hub genes. Finally, the mRNA levels of these Hub genes were quantitatively analyzed by RT-qPCR. Results We screened 181 uDEGs and 64 dDEGs, and the analysis results indicated that uDEGs and dDEGs were respectively enriched in cell component (CC), biological process (BP) and molecular functional (MF), and they were closely related to multiple signal pathways. Then, the top 25 genes with high degree score in DEGs were selected. And 8 genes could predict the poor prognosis of PAAD. RT-qPCR showed that these genes were differentially expressed in PAAD tissues. Conclusion MMP14, MMP1, MET, PLAU, ITGA2, KRT19, COL12A1 and ITGA3 are the key genes associated with the progress of pancreatic cancer.Key words: Bioinformatics analysis; Differentially expressed genes (DEGs); Pancreatic adenocarcinoma 摘 要：目的 筛选鉴定胰腺癌进展过程的关键基因和途径，并进行综合分析。
小学下册A卷英语第5单元期末试卷
小学下册英语第5单元期末试卷英语试题一、综合题(本题有100小题，每小题1分，共100分.每小题不选、错误，均不给分)1.The elephant is very ________.2.This boy, ______ (这个男孩), enjoys playing chess with friends.3.I enjoy playing with my ______. (我喜欢和我的______一起玩。
)4.The __________ (历史的影响) shapes perceptions.5.My brother collects ____ (coins) from around the world.6.The _______ (Vietnam War) involved North and South Vietnam with US involvement.7.What is the main source of light during the day?A. MoonB. StarsC. SunD. Lamp8.What do you call the process by which plants lose water?A. PhotosynthesisB. TranspirationC. RespirationD. GerminationB9.Ancient civilizations often built ________ for religious purposes.10.aust was a tragic event during __________ (二战). The Holo11.________ (植物保护倡议) promote awareness.12.My friend has a pet ______ (兔子) named Fluffy.13.He is a _____ (作家) who writes thrillers.14.听录音,按听到的顺序给下列图画标上正确的序号。
A_booming_field_of_large_animal_model_research
A booming field of large animal model researchAnimal models are integral to the study of fundamental biological processes and the etiology of human diseases.Small animal models, especially those involving mice, have yielded abundant and significant insights, greatly enhancing our understanding of biological phenomena and disease mechanisms. The preference for small animal models is primarily due to their ease of use and the availability of well-established genetic manipulation tools. The extensive use of genetically modified small animals has led to remarkable advancements in biomedical research. Nevertheless, it is important to recognize the substantial disparities in genomes,anatomy, and physiology between humans and small animals.As a result, there is an increasing awareness of the need to utilize large animal models that more closely resemble humans. Such models are essential for enhancing our understanding of crucial biological issues as well as the pathogenesis of diseases.This special column about “Large Animal Model Research ”,together with a previous review in this journal (Rahman et al.,2023), focusing specifically on large animal models and highlighting the use of various large animal models, including rabbits, pigs, and non-human primates (NHPs), in addressing key biological and medical issues. Incorporating larger animals into research not only substantially enhances the variety of research tools and methodologies available, but also enables the exploration of long-standing issues that have proven challenging using small animals, given their species-specific differences.Among the abovementioned large animals, rabbits are comparatively less commonly employed as a model system for research. However, the comprehensive review by Han et al. (2024b) offers valuable insights into the utilization of rabbit models in the field of biomedical research. Rabbits represent a cost-effective advantage over larger animals,owing to their ease of handling and rapid reproductive capacity, while also exhibiting larger body sizes and longer lifespans than rodents. These unique attributes, coupled with advancements in genetic modification techniques, such as CRISPR/Cas9 gene targeting, have led to the establishment of various rabbit models for biomedical research. Given that their lipid metabolic profiles and immune responses are more similar to humans than to rodents, rabbits are considered suitable for modeling cardiovascular and immune-related diseases. Furthermore, with their extensive involvement in commercial antibody production, genetically modified rabbits are an asset in the development of antibody-based drugs andimmunotherapeutic agents. Although the generation of rabbit models dedicated to the exploration of neurological disorders has been relatively limited, a recent study using base editing technology to modify the amyotrophic lateral sclerosis (ALS)gene SOD1 successfully produced a rabbit model exhibiting ALS phenotypes (Zhang et al., 2023). Thus, this study underscores the potential of rabbits in investigating neurological disorders.Regarding neurological disorder research, NHPs play an indispensable role due to their high similarity to humans in terms of brain structure, function, and aging processes. In their comprehensive review, Pan et al. (2024) discuss several NHP models of neurodegenerative diseases, including Alzheimer’s disease (AD), Parkinson’s disease (PD), ALS, and Huntington’s disease (HD). The analysis of these disease models underscores the significance of employing NHPs for the investigation of the fundamental mechanisms underpinning neurodegenerative diseases.Despite their extensive use over the past few decades,genetically modified mouse models cannot fully replicate the pathological features observed in human patients. A notable discrepancy is the absence of pronounced and selective neuronal loss in most genetically modified mouse models. For instance, mouse models carrying genetic mutations associated with PD do not exhibit degeneration of dopaminergic neurons, and most mouse models of ALS lack significant cytoplasmic accumulation of TDP-43, both of which are key pathological manifestations observed in human PD and ALS, respectively. In contrast, monkeys expressing PD-associated proteins (Li et al., 2021a; Yang et al., 2019) or mutant TDP-43 (Yin et al., 2019) successfully recapitulate neurodegeneration and cytoplasmic TDP-43 accumulation,respectively.Neurodegenerative features are not limited to primates; they also occur in other types of large animals. Pigs have been utilized in the exploration of neurological disorders due to their genetic, anatomical, and physiological similarities to humans.In the review by Han et al. (2024a), pigs, NHPs, and sheep were compared for their utility in studying HD, a monogenetic disorder with a genetic mutation that can be replicated across different species. Similar to mouse models of AD and PD,mouse models carrying the HD gene also fail to exhibit the robust and overt neurodegeneration observed in afflicted patient brains. Several transgenic large animal models have been established in NHPs, pigs, and sheep. Nevertheless,these models present a spectrum of phenotypic variations,and the progression and severity of disease differ significantly.Given that animal behaviors and phenotypes are strongly influenced by transgene copy number and expression levels,ideal animal models are those that can faithfully recapitulate human genetic mutations and express mutant genes at the endogenous level.Pigs possess distinct genetic modification advantages compared to NHPs and sheep. Notably, somatic nuclearThis is an open-access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (/licenses/by-nc/4.0/), which permits unrestricted non-commercial use, distribution, and reproduction in any medium,provided the original work is properly cited.Copyright ©2024 Editorial Office of Zoological Research, Kunming Institute of Zoology, Chinese Academy of SciencesLi & Lai. Zool. Res. 2024, 45(2): 311−313https:///10.24272/j.issn.2095-8137.2024.018transfer, which allows for the knock-in (KI) of mutant genes, has been successfully applied in pigs to generate a HD gene KI model that recapitulates selective neuronal loss and motor function deficiency, two crucial pathological features of HD. This HD KI pig model has been effectively employed in gene therapy research to reduce HD pathology and symptoms (Yan et al., 2023), providing further evidence for the importance of large animal models in the investigation of neurodegenerative diseases.Pigs are also extensively used in other biomedical research fields, including circulatory system diseases, organ transplantation, diabetes, skin diseases, and tumors. Porcine models also show promise for studying inherited hearing loss. Notably, pigs have a hearing range comparable to that of humans and naturally undergo age-related hearing loss, a phenomenon largely attributed to the remarkable similarity in both the structure and function of their ears to those of humans. Genetic modification techniques, such as CRISPR/Cas9 gene editing and somatic nuclear transfer, have enabled the creation of single genetic mutations in pig models, facilitating the study of disease pathogenesis in monogenic hearing loss. For example, Wang et al. (2024) discussed the use of pig models in studying hearing loss-related diseases, with notable implications for other monogenic diseases.The four research articles published in this special issue also highlight the significance of large animal models in research. Li et al. (2024a) reported on mitochondrial replacement in cynomolgus monkeys (Macaca fascicularis) through female pronuclear transfer. Their findings suggest that pronuclear transfer holds great potential in reducing the risk of inherited mitochondrial DNA (mtDNA) diseases, thereby establishing the applicability of non-human models in investigating diseases related to mtDNA defects. Li et al. (2024b) developed cynomolgus monkey organoids to study neural tube defects (NTDs), which can also arise from loss-of-function mutations in the SHROOM3 gene. Their findings indicate that in vitro models using NHP-derived organoids can be employed to investigate the pathogenesis of significant human diseases.The rationale for using NHPs in research is also based on species-dependent gene expression patterns. Chen et al. (2024) presented intriguing findings regarding the expression of PINK1, which is associated with PD, in mice, pigs, and monkeys. Using multiple antibodies and PINK1 knockout animal models, they discovered that the PINK1 protein, rather than PINK1 mRNA, is detectably and exclusively expressed in the primate brain. These findings explain why PINK1 knockout leads to neurodegeneration in monkeys, but not in mice and pigs, suggesting that a possible higher abundance of endogenous PINK1 in primate brains contributes to neuronal survival. Their results also imply that species-dependent transcriptional and translational regulations contribute to species-specific pathology. Therefore, utilizing animal models that more closely resemble humans provides better prospects for uncovering the molecular mechanisms underlying primate-specific gene expression. Reinforcing this idea, Mao et al. (2024) conducted a comprehensive analysis of the transcriptomes within macaque species and between macaques and humans, highlighting the high conservation of tissue-specific genes and the value of macaques as biological models for investigating human diseases. Additionally, the identification of a cynomolgus monkey with naturally occurring PD further supports the suitability of NHPs as ideal models for PD research (Li et al., 2021b).Despite the significant contributions and advancements in large animal models for biomedical research, notable limitations and challenges exist. The utilization of large animals necessitates stricter regulation and control due to ethical concerns arising from their high cognitive capacities and close resemblance to humans. Additionally, the substantial costs and extended breeding periods associated with maintaining large animals pose considerable challenges in expanding the scope of studies using these models. Undoubtedly, small animal models, particularly rodents, are likely to remain fundamental in biological and biomedical research, continuing to play a critical role in enhancing our understanding of disease pathogenesis and in developing therapeutic strategies for human diseases. However, in scenarios where small animal models have failed to replicate important pathological events observed in humans, it is worth considering large animal alternatives to unravel disease mechanisms that may not be evident in small animals. Furthermore, large animal models can facilitate the validation of specific therapeutic targets, potentially reducing the failure rates of clinical trials for therapeutics initially developed using small animal models. The integration of large animal models into research endeavors promises to deepen our understanding of complex biological processes and improve the translation of preclinical findings into clinical practice.Xiao-Jiang Li1,2,*, Liangxue Lai3,*1 Guangdong Key Laboratory of Non-human Primate Research,Key Laboratory of CNS Regeneration (Ministry of Education), GHM Institute of CNS Regeneration, Jinan University,Guangzhou, Guangdong 510632, China2 State Key Laboratory of Bioactive Molecules and Druggability Assessment, Jinan University, Guangzhou, Guangdong 510632,China3 CAS Key Laboratory of Regenerative Biology, GuangdongProvincial Key Laboratory of Stem Cell and RegenerativeMedicine, Guangzhou Institutes of Biomedicine and Health, Chinese Academy of Sciences, Guangzhou, Guangdong 510530,China*Corresponding authors, E-mail: **************.cn;********************.cnREFERENCESChen XS, Han R, Liu YT, et al. 2024. Comparative analysis of primate and pig cells reveals primate-specific PINK1 expression and phosphorylation. Zoological Research, 45(2): 242−252.Han B, Liang W, Li XJ, et al. 2024a. Large animal models for Huntington’s disease research. Zoological Research, 45(2): 275−283.Han Y, Zhou J, Zhang R, et al. 2024b. Genome-edited rabbits: Unleashing the potential of a promising experimental animal model across diverse diseases. Zoological Research, 45(2): 253−262.Li CY, Liu XC, Li YZ, et al. 2024a. Generation of mitochondrial replacement monkeys by female pronucleus transfer. Zoological Research, 45(2): 292−298.Li H, Wu SH, Ma X, et al. 2021a. Co-editing PINK1 and DJ-1 genes via adeno-associated virus-delivered CRISPR/Cas9 system in adult monkey brain elicits classical parkinsonian phenotype. Neuroscience Bulletin, 37(9): 1271−1288.Li H, Yao YG, Hu XT. 2021b. Biological implications and limitations of a312 cynomolgus monkey with naturally occurring Parkinson's disease. Zoological Research, 42(2): 138−140.Li P, Zhang T, Wu R, et al. 2024b. Loss of SHROOM3 affects neuroepithelial cell shape through regulating cytoskeleton proteins in cynomolgus monkey organoids. Zoological Research, 45(2): 233−241.Mao YX, Li Y, Yang Z, et al. 2024. Comparative transcriptome analysis between rhesus macaques (Macaca mulatta) and crab-eating macaques (Macaca fascicularis). Zoological Research, 45(2): 299−310.Rahman A, Li Y, Chan TK, et al. 2023. Large animal models of cardiac ischemia-reperfusion injury: Where are we now?. Zoological Research, 44(3): 591−603.Pan MT, Zhang H, Li XJ, et al. 2024. Genetically modified non-human primate models for research on neurodegenerative diseases. Zoological Research, 45(2): 263−274.Wang X, Liu TX, Zhang Y, et al. 2024. Genetically modified pigs: Emerging animal models for hereditary hearing loss. Zoological Research, 45(2): 284−291.Yan S, Zheng X, Lin YQ, et al. 2023. Cas9-mediated replacement of expanded CAG repeats in a pig model of Huntington's disease. Nature Biomedical Engineering, 7(5): 629−646.Yang WL, Liu YB, Tu ZC, et al. 2019. CRISPR/Cas9-mediated PINK1 deletion leads to neurodegeneration in rhesus monkeys. Cell Research, 29(4): 334−336.Yin P, Guo XY, Yang WL, et al. 2019. Caspase-4 mediates cytoplasmic accumulation of TDP-43 in the primate brains. Acta Neuropathologica, 137(6): 919−937.Zhang ZT, Wu XY, Yang J, et al. 2023. Highly efficient base editing in rabbit by using near-PAMless engineered CRISPR/Cas9 variants. Science China Life Sciences, 66(3): 635−638.Zoological Research 45(2): 311−313, 2024 313。
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Introduction
Chitin, a linear polymer of β-(1, 4)-linked N-acetylglucosamines, is one of the most crucial component in the cuticle, cuticular lining of the foregut, hindgut, trachea, and peritrophic membrane (PM) in insects [1-2], and widely distributed in fungi, nematodes and other arthropods [3]. During each molting cycle in insect development, part of the old cuticle is digested, while new chitin is synthesized and deposited [4]. Chitin synthase (CHS) is a crucial enzyme involved in the final step of the chitin biosynthetic pathway. CHSs are large proteins embedded in the plasma membrane and belong to family 2 of the glycosyltransferases, which catalyze the transfer of sugar moieties from activated sugar donors to specific acceptors resulting in a glycosidic bond [5]. Based on amino acid sequence similarity, distribution, and function, insect chitin synthase occurs in two types: CHS1 and CHS2 (also known as CHS-A
Received: 2013.02.02; Accepted: 2013.03.31; Publisho alternative splicing variants of chitin synthase 1 gene (BdCHS1) were cloned and characterized from the oriental fruit fly, Bactrocera dorsalis (Hendel). The cDNA of both variants (BdCHS1a and BdCHS1b) consisted of 5,552 nucleotides (nt), with an open reading frame (ORF) of 4,776 nt, encoding a protein of 1,592 amino acid residues, plus 685- and 88-nt of 5′- and 3′-noncoding regions, respectively. The alternative splicing site was located between positions 3,784–3,960 and formed a pair of mutually exclusive exons (a/b) that were same in size (177 nt), but showed only 65% identity at the nucleotide level. During B. dorsalis growth and development, BdCHS1 and BdCHS1a were both mainly expressed during the larval–pupal and pupal–adult transitions, while BdCHS1b was mainly expressed during pupal–adult metamorphosis and in the middle of the pupal stage. BdCHS1a was predominately expressed in the integument whereas BdCHS1b was mainly expressed in the trachea. The 20-hydroxyecdysone (20E) induced the expression of BdCHS1 and its variants. Injection of dsRNA of BdCHS1, BdCHS1a, and BdCHS1b into third-instar larvae significantly reduced the expression levels of the corresponding variants, generated phenotypic defects, and killed most of the treated larvae. Furthermore, silencing of BdCHS1 and BdCHS1a had a similar result in that the larva was trapped in old cuticle and died without tanning completely, while silencing of BdCHS1b has no effect on insect morphology. These results demonstrated that BdCHS1 plays an important role in the larval–pupal transition and the expression of BdCHS1 in B. dorsalis is regulated by 20E.
Int. J. Biol. Sci. 2013, Vol. 9
331
Ivyspring
International Publisher
International Journal of Biological Sciences
2013; 9(4):331-342. doi: 10.7150/ijbs.6022
© Ivyspring International Publisher. This is an open-access article distributed under the terms of the Creative Commons License (/ licenses/by-nc-nd/3.0/). Reproduction is permitted for personal, noncommercial use, provided that the article is in whole, unmodified, and properly cited.
Int. J. Biol. Sci. 2013, Vol. 9
and CHS-B) [6-9]. These two genes are closely related, but belong to different phylogenetic clades [2]. CHS1 and CHS2 have different functions during insect growth and development. CHS1 is responsible for production of chitin required in cuticle and cuticular lining of the foregut, hindgut and trachea, whereas CHS2 specializes in chitin synthesis in the PM [1, 10]. However, only CHS1 has been found in hemipterans such as pea aphids, triatomine bugs, and brown planthoppers, while the CHS2 gene was absent, consistent with the lack of PM in these insects [11, 12]. The hemipterans may have lost CHS2 during their evolution. In addition, insect CHS1 consists of two alternative splicing variants, CHS1a and CHS1b, which differ in a 177-nt region that encodes 59 amino acids forming the predicted second to last transmembrane helix [7, 13-15]. No alternative splicing variants have been reported for CHS2 in insects [3, 16]. Due to the fact that chitin is absent in plants and vertebrates [5], CHS represents a potential target for selective insecticidal agents. Many insect growth regulators, such as diflubenzuron and chlorfluazuron, have been reported to disrupt chitin synthesis in insects [17, 18]. In addition, 20-hydroxyecdysone (20E) and the ecdysone agonist RH5992 may be able to interfere with insect cuticle formation by inhibiting CHS1 [19]. In a recent study, serious cuticular malformations were observed and CHS1 expression was up-regulated as a result of 20E treatment of a lepidopteran [20]. There have been no reports on the effects of 20E on CHS1 in tephritid fruit flies to date. The oriental fruit fly, Bactrocera dorsalis (Hendel) (Diptera: Tephritidae), is one of the most economically important pests in tropical and subtropical areas of the world, where it causes serious damage to fruit production [21, 22]. Females typically oviposit in fruit, and the developing larvae tunnel through the fleshy mesocarp on which they feed, causing fruit damage and drop [23]. Current control techniques mainly rely on spraying chemical insecticides. Because of the high economic and environmental costs of chemical control, together with the evolution of insecticide-resistant populations, there is an urgent need to develop new pest management strategies. In recent years, RNA interference (RNAi)-based technology has shown great potential in controlling insect pests by silencing vital genes [24, 25]. RNAi has also been successfully used to investigate the developmental function of CHS1 in holometabolous and hemimetabolous insects, such as Tribolium castaneum (Coleoptera) [26], Drosophila melanogaster (Diptera) [27, 28], Spodoptera exigua (Lepidoptera) [29], Ostrinia furnacalis (Lepidoptera) [30], Locusta migratoria manilensis (Orthoptera) [14], and Nilaparvata lugens (Hemiptera) [11]. These results showed that CHS1 is required for sur-