a novel approach for transcription factor analysis using selex with high-throughput sequencing (tfast)一种新型转录因子分析方法使用高通量测序的selex(tfast).pdfVIP

A Novel Approach for Transcription Factor Analysis Using SELEX with High-Throughput Sequencing (TFAST) 1,2 2 1,2 Daniel J. Reiss , Frederick M. Howard , Harry L. T. Mobley * 1 Department of Microbiology and Immunology, University of Michigan, Ann Arbor, Michigan, United States of America, 2 University of Michigan Medical School, Ann Arbor, Michigan, United States of America Abstract Background: In previous work, we designed a modified aptamer-free SELEX-seq protocol (afSELEX-seq) for the discovery of transcription factor binding sites. Here, we present original software, TFAST, designed to analyze afSELEX-seq data, validated against our previously generated afSELEX-seq dataset and a model dataset. TFAST is designed with a simple graphical interface (Java) so that it can be installed and executed without extensive expertise in bioinformatics. TFAST completes analysis within minutes on most personal computers. Methodology: Once afSELEX-seq data are aligned to a target genome, TFAST identifies peaks and, uniquely, compares peak characteristics between cycles. TFAST generates a hierarchical report of graded peaks, their associated genomic sequences, binding site length predictions, and dummy sequences. Principal Findings: Including additional cycles of afSELEX-seq improved TFAST’s ability to selectively identify peaks, leading to 7,274, 4,255, and 2,628 peaks identified in two-, three-, and four-cycle afSELEX-seq. Inter-round analysis by TFAST identified 457 peaks as the strongest candidates for true binding sites. Separating peaks by TFAST into classes of worst, second-best and best candidate peaks revealed a trend of increasing significance (e-values 4.5 61012, 2.9 610246, and 1.2610273) and