Browsing by Author "Huang, Tim H."

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    A New Method for Stranded Whole Transcriptome RNA-seq
    (Elsevier, 2013) Miller, David F. B.; Yan, Pearlly S.; Buechlein, Aaron; Rodriguez, Benjamin A.; Yilmaz, Ayse S.; Goel, Shokhi; Lin, Hai; Collins-Burow, Bridgette; Rhodes, Lyndsay V.; Braun, Chris; Pradeep, Sunila; Rupaimoole, Rajesha; Dalkilic, Mehmet; Sood, Anil K.; Burow, Matthew E.; Tang, Haixu; Huang, Tim H.; Liu, Yunlong; Rusch, Douglas B.; Nephew, Kenneth P.; Cellular and Integrative Physiology, School of Medicine
    This report describes an improved protocol to generate stranded, barcoded RNA-seq libraries to capture the whole transcriptome. By optimizing the use of duplex specific nuclease (DSN) to remove ribosomal RNA reads from stranded barcoded libraries, we demonstrate improved efficiency of multiplexed next generation sequencing (NGS). This approach detects expression profiles of all RNA types, including miRNA (microRNA), piRNA (Piwi-interacting RNA), snoRNA (small nucleolar RNA), lincRNA (long non-coding RNA), mtRNA (mitochondrial RNA) and mRNA (messenger RNA) without the use of gel electrophoresis. The improved protocol generates high quality data that can be used to identify differential expression in known and novel coding and non-coding transcripts, splice variants, mitochondrial genes and SNPs (single nucleotide polymorphisms).
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    Estrogen induces global reorganization of chromatin structure in human breast cancer cells
    (PLoS, 2014-12-03) Mourad, Raphael; Hsu, Pei-Yin; Juran, Liran; Shen, Changyu; Koneru, Prasad; Lin, Hai; Liu, Yunlong; Nephew, Kenneth; Huang, Tim H.; Li, Lang; Department of Medical and Molecular Genetics, IU School of Medicine
    In the cell nucleus, each chromosome is confined to a chromosome territory. This spatial organization of chromosomes plays a crucial role in gene regulation and genome stability. An additional level of organization has been discovered at the chromosome scale: the spatial segregation into open and closed chromatins to form two genome-wide compartments. Although considerable progress has been made in our knowledge of chromatin organization, a fundamental issue remains the understanding of its dynamics, especially in cancer. To address this issue, we performed genome-wide mapping of chromatin interactions (Hi-C) over the time after estrogen stimulation of breast cancer cells. To biologically interpret these interactions, we integrated with estrogen receptor α (ERα) binding events, gene expression and epigenetic marks. We show that gene-rich chromosomes as well as areas of open and highly transcribed chromatins are rearranged to greater spatial proximity, thus enabling genes to share transcriptional machinery and regulatory elements. At a smaller scale, differentially interacting loci are enriched for cancer proliferation and estrogen-related genes. Moreover, these loci are correlated with higher ERα binding events and gene expression. Taken together these results reveal the role of a hormone--estrogen--on genome organization, and its effect on gene regulation in cancer.