小分子RNA- sma rna
小分子RNA--small RNA
什么是干扰小RNA? mirnA(微小RNA): MicroRNAs (miRNAs)即微小RNA是在所有真核类中发 的平均长度为22nt的小分子RNA. mi rnas产生于 mRNA pre-mRNA),可与靶mRNA配对并抑制mRNA的翻译 或促使其降解,具有转录后调控基因表达的功能人 类基因组中约有1000种mRNA,可影响60%的基因的表 达与调控 SiRNA(小RNA): mall interfering RNA (siRNA), sometimes known as short interfering rna or silencing rna, is a class of double-stranded RNA molecules, 20-25 nucleotides in length, that play a variety of roles in biology. Most notably, sirNa is involved in the rna interference(RNAi) pathway, where it interferes with the expression of a specific gene. See:http:/en.wikipedia.org/wiki/smallinterferingrna
什么是干扰小RNA? miRNA(微小RNA): MicroRNAs (miRNAs) 即微小RNA是在所有真核类中发现 的平均长度为22nt的小分子RNA. miRNAs 产生于前体 mRNA(pre-mRNA), 可与靶mRNA配对并抑制mRNA的翻译 或促使其降解, 具有转录后调控基因表达的功能. 人 类基因组中约有1000种miRNA, 可影响60%的基因的表 达与调控. siRNA(小RNA): Small interfering RNA (siRNA), sometimes known as short interfering RNA or silencing RNA, is a class of double-stranded RNA molecules, 20-25 nucleotides in length, that play a variety of roles in biology. Most notably, siRNA is involved in the RNA interference (RNAi) pathway, where it interferes with the expression of a specific gene. See: http://en.wikipedia.org/wiki/Small_interfering_RNA
干扰小RNA的种类 有4大类干扰小RNA( (small interfering RNa) D)miRNA, Micro-interfering rna 2)SiRNA, Short-interfering RNA 3) piRNA, Piwi-interacting RNa,仅限动物 4)2 IU RNA,线虫
干扰小RNA的种类 有4大类干扰小RNA (small interfering RNA): 1) miRNA, Micro-interfering RNA 2) siRNA, Short-interfering RNA 3) piRNA, Piwi-interacting RNA, 仅限动物 4) 21U RNA, 线虫
microrNa
microRNA
microrNA是如何发现的? 殊途同归 1)植物转基因沉默 2)真菌转基因沉默 3)线虫RNA注射实验 4)线虫发育突变研究 4个独立的研究均发现RNA诱导 内源基因沉默
microRNA是如何发现的? 殊途同归: 1) 植物转基因沉默 2) 真菌转基因沉默 3) 线虫RNA注射实验 4) 线虫发育突变研究 4个独立的研究均发现RNA诱导 内源基因沉默
植物转基因 Dr Marjori Matzke Dr. antonius 沉默 Matzke Wild Flowers of chalcone synthase V26 petunia ilenced transformants 1989年 Matzke等为了加深矮牵牛花瓣的颜色将控制紫色花的基因导入矮牵牛细胞, 并获得再生植株未曾预料的是在转基因阳性植株中出现完全为白色花瓣的植株 深入研究发现,导入矮牵牛的转基因含有反向尾-尾重复的结构产生正义和反义 基因转录产物正义和反义基因转录产物形成 dsrNA,导致转基因和内源基因的沉 默使原本产生紫色花色的花瓣变成白色.这是首次报道正反义双链RNA导致基因 Y. Posttranscriptional gene silencing(PTGS)in plants
植物转基因 沉默 1989年Matzke等为了加深矮牵牛花瓣的颜色将控制紫色花的基因导入矮牵牛细胞, 并获得再生植株.未曾预料的是,在转基因阳性植株中出现完全为白色花瓣的植株. 深入研究发现, 导入矮牵牛的转基因含有反向尾--尾重复的结构. 产生正义和反义 基因转录产物.正义和反义基因转录产物形成dsRNA, 导致转基因和内源基因的沉 默,使原本产生紫色花色的花瓣变成白色. 这是首次报道正反义双链RNA导致基因 沉默. Posttranscriptional gene silencing (PTGS) in plants. Dr. Marjori Matzke Dr. Antonius Matzke
真菌转基因诱导的内源基因沉默 (PTGS) Isolation of quelling-defective(gde) mutants impaired in posttrans criptional transgene-induced gene silencing in Neurospora crassa, Proc. Natl. Acad. Sci USA 94(1997: These quelling-defective mutants(gde) belonging to three complementation groups have provided insights into the mechanism of posttranscriptional gene silencing in N. crassa. The recessive nature of the gde mutations indicates that the encoded gene products act in trans. we show that when gde genes are mutated in a transgenic-induced silenced strain containing many copies of the transgene the expression of the endogenous gene is maintained despite the presence of transgene sense RNA, the molecule proposed to trigger quelling
真菌转基因诱导的内源基因沉默 (PTGS) Isolation of quelling-defective (qde) mutants impaired in posttranscriptional transgene-induced gene silencing in Neurospora crassa, Proc. Natl. Acad. Sci. USA 94 (1997): These quelling-defective mutants (qde) belonging to three complementation groups have provided insights into the mechanism of posttranscriptional gene silencing in N. crassa. The recessive nature of the qde mutations indicates that the encoded gene products act in trans. We show that when qde genes are mutated in a transgenic-induced silenced strain containing many copies of the transgene, the expression of the endogenous gene is maintained despite the presence of transgene sense RNA, the molecule proposed to trigger quelling
线虫双链RNA分子导致的基因沉默 1995年Come大学的研究人员uo和 Kemphues在研究阻断秀 丽新小杆线虫中的ar基因时,利用反义RNA( antisense RNA)技术特异性地阻断par-l基因的表达,同时在对照实验 注射正义RNA( sense rNA)观察基因表达是否增强。但结 果是二者都同样地切断了par-1基因的表达途径,这与传统 的反义RNA功能解释自相矛盾,作者对此未给出合理解释 1998年2月,Fre和Meo首次揭开谜底。他们证实,Guo等报 道的正义RNA抑制基因表达的现象是由于实验中污染了微量 双链RNA( Double rna, dsRNA)所致。他们将体外转录的单 链RNA纯化后注射线虫,发现基因抑制效果士分微弱。而经 过纯化的双链RNA却正好相反,能够高效特异性阻断靶基因 的表达,其效率高于单链反义RNA约2个数量级。 Fire等对 dsRNA调控内源基因表达的现象创造了一个名词: rna interference,简称RNAi,即RNA干扰 See: Fire A, (1998). "Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans". Nature 391: 806-11
线虫双链RNA分子导致的基因沉默 1995年Cornell大学的研究人员Guo和Kemphues在研究阻断秀 丽新小杆线虫中的par-1基因时，利用反义RNA (antisense RNA)技术特异性地阻断par-1基因的表达，同时在对照实验 注射正义RNA（sense RNA）观察基因表达是否增强。但结 果是二者都同样地切断了par-1 基因的表达途径,这与传统 的反义RNA功能解释自相矛盾，作者对此未给出合理解释。 1998年2月，Fire和Mello首次揭开谜底。他们证实，Guo等报 道的正义RNA抑制基因表达的现象是由于实验中污染了微量 双链RNA (Double RNA, dsRNA)所致。他们将体外转录的单 链RNA纯化后注射线虫，发现基因抑制效果十分微弱。而经 过纯化的双链RNA却正好相反，能够高效特异性阻断靶基因 的表达，其效率高于单链反义RNA约2个数量级。 Fire等对dsRNA调控内源基因表达的现象创造了一个名词: RNA interference, 简称RNAi, 即RNA干扰. See: Fire A, (1998). "Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans". Nature 391:806–11
以线虫为模式生物研究 双链RNA对基因表达的 干扰 Nature391:806-811 2006 Dr Fire uses c elegans as a godel in RNAi work 1998 年生 Potent and specific 理与 genetic interference by Andrew Fire double-stranded rain 医学 Caenorhabditis elegans 诺贝 Andrew Fire, siQun Xu, Mary K. Montgomery 尔奖 Steven A Kostas*t, Samuel E Driver* Craig C Mellot Carnegie Institution of Washington, Department of Embryology 115 West University Parkway Baltimore, Maryland 21210, USA t Biology Graduate Program, Johns Hopkins University, 3400 North Charles Street, Baltimore, Maryland 21218, USA t Program in Molecular Medicine, Department of Cell Biolog University of Massachusetts Cancer Center, Tivo Biotech Suite 213, Craig C. Mello 373 Plantation Street, Worcester, Massachusetts 01605, USA
2006 年生 理与 医学 诺贝 尔奖
Effects of mex-3 RNA Negative control showing lack of staining in the interference on /eve/s of the absence of the hybridization endogenous MRNA probe. b, Embryo from uninjected parent(showing normal pattern of endogenous mex-3 RNA20 Embryo from a parent injected with purified mex 3B antisense RNA. These embryos(and the parent animals) retain the mex-3 MRNA, although levels may be somewhat less than wild type d, Embryo from a parent injected with dsRNA corresponding to MEX-3 is a Kh domain protein that mex-3B: no mex-3 RNA is regulates blastomere(卵裂球) identity detected. each embryo is in early C elegans embryos approximately 50 mm in length
Effects of mex-3 RNA interference on levels of the endogenous mRNA MEX-3 is a KH domain protein that regulates blastomere (卵裂球) identity in early C. elegans embryos a, Negative control showing lack of staining in the absence of the hybridization probe. b, Embryo from uninjected parent (showing normal pattern of endogenous mex-3 RNA20). c, Embryo from a parent injected with purified mex- 3B antisense RNA. These embryos (and the parent animals) retain the mex-3 mRNA, although levels may be somewhat less than wild type. d, Embryo from a parent injected with dsRNA corresponding to mex-3B; no mex-3 RNA is detected. Each embryo is approximately 50 mm in length