CloningofsmallRNAswith5’phosphateand3’OHends2
3’ Adaptor Ligation and PurificationHeat shock the RNA by putting at 90°C for 30 seconds. Snap cool on ice.Set up the 3’Adaptor ligation reaction in a 1.5 ml siliconized tube: Purified 5’ ligation product from step 2.15 6.4µl3’ RNA adaptor (10µM) ......阅读全文
Cloning－of－small－RNAs－with－5’－phosphate－and－3’－OH－ends2
3’ Adaptor Ligation and PurificationHeat shock the RNA by putting at 90°C for 30 seconds. Snap cool on ice.Set up the 3’Adaptor ligation reaction in a
Cloning－of－small－RNAs－with－5’－phosphate－and－3’－OH－ends3
Load your precipitated PCR samples into 2 consecutive lanes so as not to overload the lanes. For each different sample, I would run a separate ladder
Cloning－of－small－RNAs－with－5’－phosphate－and－3’－OH－ends01
IntroductionThe following protocol describes a procedure for the purification and cloning of miRNAs and other small RNAs in the 20-30 nucleotide size
RNAi制备小分子干扰RNA（small－interfering－RNAs，siRNAs)的方法2
越来越多的研究人员开始采用小分子干扰RNA（small interfering RNAs，siRNAs)来抑制特定的哺乳动物基因表达。siRNA是一种短片断双链RNA分子，能够以同源互补序列的mRNA为靶目标降解特定的 mRNA，这个过程就是RNA干扰途径（RNA interference
RNAi制备小分子干扰RNA（small－interfering－RNAs，siRNAs)的方法1
最适用于：快速而经济地研究某个基因功能缺失的表型不适用于：长时间的研究项目，或者是需要一个特定的siRNA进行研究，特别是基因治疗体内表达前面的3种方法主要都是体外制备siRNAs，并且需要专门的RNA转染试剂将siRNAs转到细胞内。而采用siRNA表达载体和基于PCR的表达框架则属于：从转染到细
RNAi术语表
RNAi GlossaryDicer - Dicer is a member of the RNase III family of nucleases that specifically cleave double-stranded RNAs. Dicer processes long dsRNA
Phosphate－Assay
1. Make standards using sodium phosphate at the following uM concentrations: 0, 2, 5, 7, 10, 20, 40, 60, and 80. Use the screw top glass tubes.2. Dry
RNAi相关的名词解释[英文]
Argonaute - A family of proteins containing multiple domains and involved in RNA interference (RNAi). Argonatue is the main component of RNAi effector
Fusion－and－Cloning
Author: Nanci DonackiSource: Contributed by Nanci DonackiAbstract: Procedure for establishing hybridoma in one stepReagents(StemCell Technologies, Inc
Fusion－and－Cloning
ReagentsMedium A - Pre-fusion Medium and Hybridoma Expansion MediumMedium B - Fusion Medium Medium C - Hybridoma Recovery MediumMedium D - Hybridoma S
DNA转化实验指导2
1B. Cloning 1. A caveat on dephosphorylation: the most common reason for failure to obtain colonies is a result of adding too much BAP or CIP to
Cloning－PCR－products－using－TA－vectors
Cloning PCR products using TA vectorsby Paul N. Hengen, Ph.D. *Methods and reagents is a unique monthly column that highlights current discussions in
General－Cloning－Protocols
Large Scale Preps: (See Large scale plsasmid prep protocol for more details)Cultures: Inoculate a 5 mL LB/Amp (50 - 100 µg/mL) culture in early a.m. w
PCR基本实验方法（五）
Cloning PCR ProductsT-A Cloning Strategy: Taq and other polymerases seem to have a terminal transferase activity which results in the non-templated ad
PCR基本实验方法（五）
Cloning PCR ProductsT-A Cloning Strategy: Taq and other polymerases seem to have a terminal transferase activity which results in the non-templated ad
Simplified－Calcium－Phosphate－Coprecipitation
Materials2x HEPES: 8 g NaCl, 0.105g NaHPO4, 6.5 g HEPES, H2O to 500 mL, pH 6.95 to 7.10 (try a range)2M CaCl2: store in aliquots at -20°CDNA 4 mg/mLPr
Phosphate－(Sodium)－buffer－Chart
Phosphate (Sodium) buffer ChartStock solution A2 M monobasic sodium phosphate, monohydrate (276g/L)Stock solution B2 M dibasic sodium phosphate (284 g
Phosphate－Assay－by－Suprya－Jaydev
ReagentsAshing buffer:10 g Mg(NO3)2 100 ml EtOH1.5 N HCl stock:119.7 ml concentrated HCl (11.6M @ 36% by weight)880.3 ml H2O1 N Sulfuric acid stock:28
Cloning－by－Limiting－Dilution－of－Hybridoma
Author: Nanci DonackiSource: Contributed by Nanci DonackiDate Added: Tue May 14 2002Date Modified: Tue Apr 27 2004MaterialsDMEM, high glucose (Life Te
基因cloning经验指南
我们克隆基因的时候，往往可以通过一些途径（如pcr,EST库或者文库筛选）得到基因的部分片段。然后通过3'race 和 5'race 方法往两端延伸。有时会出现无法延伸的状况，如果你超作没有失误的话，这时候很有可能是你模板GC含量过高的缘故，普通pcr 是没有办法延伸的，可以用扩高g
Genomic－Cloning－Technical－Manual
Genomic Cloning Technical ManualAn optimal strategy for genomic cloning should meet three requirements: 1) a maximum number of recombinants should be
Oxidative－reactions－of－the－pentose－phosphate－pathway
One form of chemical energy used to drive biosynthetic reactions forward is the reducing power of the energy carrier NADPH. NADPH is essential to driv
RNAi－protocol
siRNA protocolsOur current strategy with siRNA is to synthesis relatively small amounts enzymatically and use these to test for efficiency by western
NAi－protocol
siRNA protocolsOur current strategy with siRNA is to synthesis relatively small amounts enzymatically and use these to test for efficiency by western
鹌鹑PIWI基因的克隆及其结合小RNA的鉴定
PIWI 蛋白能通过与多种内源非编码小RNA结合在表观水平和转录后水平调控基因的表达。然而在家禽中，PIWI蛋白的生物学功能及其结合小RNA的作用机制尚不清楚。扬州大学动物科学与技术学院陈国宏教授课题组对该作用机理进行了研究*，研究成果发表在2012年12月刊的PLoS ONE上。PIWI 基因
“电子”基因克隆－（sillcon－cloning）
利用计算机来协助克隆 基因，称为“电子”基因克隆 （sillcon cloning），是与定位克隆 、定位候选克隆 策略并列的方法之一，即采用生物信息学的方法延伸EST序列，以获得基因部分乃至全长的cDNA序列。EST数据库的迅速扩张，已经并将继续导致识别与克隆 新基因策略发生革命性变化。1
磷酸缓冲液（phosphate－buffer）
按照下表所给定的体积，混合1 mol/L 的磷酸二氢钠（单碱）和1mol/L 磷酸氢二钠（双碱）贮液，获得所需pH的磷酸缓冲液。配制1 mol/L 的磷酸二氢钠（NaH2PO4•H2O）贮液：溶解138g于足量水中，使终体积为1L;1mol/L 磷酸氢二钠（Na2HPO4）贮液:溶解14
Experimental－Protocol－for－cDNA－Library－Construction
Experimental Protocol for cDNA Library ConstructionIdentify appropriate celltype over-expressing corresponding gene.Find out if transcription can be s
非编码small－RNA参与缺氧下的血管生成（一）
非编码RNA是近年来转录组学研究的热点，其中，long non-coding RNA（lncRNA），microRNA，circularRNA是大家研究的非常多的非编码。其实，在small non-coding RNAs世界，除了我们熟知的microRNA之外，还包括piwi-interac
Detection－of－apoptotic－process－in－situ－using－immunocytochemical
1. INTRODUCTION Apoptosis was observed from invertebrates to lower and higher verterbrates, and intervenes both in physiological and in pathological