A Method For Detecting Transcription Direction Of Gene And The Kit Development

High-throughput sequencing has identified a large number of sense-antisense transcriptional pairs,which indicates that these genes were transcribed from both directions.Recent reports have demonstrated that many antisense RNAs,especially lncRNA,can interact with the sense RNA by forming an RNA duplex.Many methods,such as RNAsequencing,Northern blotting,RNase protection assays and strand-specific PCR,can be used to detect the antisense transcript and gene transcriptional orientation.However,the applications of these methods have been constrained,to some extent,because of the high cost,difficult operation or inaccuracy,especially regarding the analysis of substantial amounts of data.Thus,we developed an easy method to detect and validate these complicated RNAs.We primarily took advantage of the strand specificity of RT-PCR and the single-strand specificity of S1 endonuclease to analyze sense and antisense transcripts.Six known genes,including ?-actin,Tsix,LXN,THRA,LCT and BRCA2 were used to establish the method.The results indicated that the method can easily distinguish sense,antisense and sense-antisense transcriptional pairs.In addition,it can be used to verify the results of high-throughput sequencing,as well as to analyze the regulatory mechanisms between RNAs.In addition,we developed the regaent kit to detect gene transcriptional orientation which is intended for research use only.