Rat ciliary neurotrophic factor in the design and validation of siRNA in vitro method.docVIP

 PAGE \* MERGEFORMAT 19 Rat ciliary neurotrophic factor in the design and validation of siRNA in vitro method On: Zheng Xiang, Li Y, Sun Yun-flower, DING Yan, Bi Wenjie, ZHOU Xue [Keywords:] RNA interference; ciliary neurotrophic factor (CNTF); rat; in vitro RNA interference (RNAi) in recent years developed a great potential for medical applications of reverse genetics technology [1]. The technology has the double-stranded interfering RNA design and synthesis, in vitro the effectiveness of sequence and in vivo experiments and other important link [2]. which is to ensure that the first two links were successfully pre-condition. and the plants and lower animals, in mammalian cells RNA interference conditions were more demanding, there are still many difficulties. For example, in vitro experiments , small interfering RNA (siRNA) design and validation must be based on the laws of a particular [3-5], and the higher requirements of the operator. Nevertheless, we continue in the experiment lessons learned, that many siRNA design Synthesis and effect of a strong validation of the operational rules to follow, with a certain experience can be more effective. Therefore, our rat CNTF (ciliary neurotrophic factor, CNTF) of RNA interference as an example, siRNA design Synthesis and in vitro validation of the whole process, I hope these experiences can provide a number of other experimental helpful information. 1 Materials and methods 1.1 rat CNTF siRNA synthesis and modification of molecular design In the PubMed home page (URL http: /pubmed/) in the cDNA sequence of rat CNTF Links Accession number “NM_013166”. Into Invitrogen’s BLOCK iTTM RNAi Designer home page, select “siRNA “design pattern (ie 19nt target sequence selection), followed by the following: the rat CNTF mRNA in the Accession number entered in the” Accession number “dialog box, check” Open reading frame (ORF) “option (expressed in open reading box to find the target sequence), select the spec