title: WUCA-Cont-29
organism: Homo sapiens
description: Protocols: We retrospectively obtained de-identified FFPE tissue from patients who underwent ileo-colic resection surgery at Barnes-Jewish Hospital, St. Louis between 2005 and 2010. CD patient surgical resections were due to complications associated with the disease, whereas control resections were due to adenoma, carcinoma or abdominal trauma. The set of hematoxylin and eosin-stained tissue sections from each resection were obtained and examined by two pathologists (T.C.L. and T.S.S.) to identify the uninvolved proximal margin (terminal ileum) of each resection. These tissue sections were then further evaluated to identify those that met the histological inclusion criteria for this study. The criteria were that the sample had at least 100 well-oriented intestinal crypts (contained crypt lumens that extended from the apex to the base of the crypt) and absent/minimal active or chronic inflammatory disease (e.g., cryptitis, crypt abscess, architectural distortion, pyloric gland metaplasia, etc.).The study protocol was approved by the Institutional Review Board of Washington University. Patients were recruited by the Washington University Digestive Diseases Research Core Center (DDRCC). Written informed consent was obtained from all study participants prior to inclusion in the study. For samples that met the histological inclusion criteria, two fresh, unstained 5-µm FFPE tissue sections on glass microscope slides were obtained. Slides were not baked after cutting. For each sample, the two tissue sections were scraped off of the slides using a clean razor and placed in 1.5 mL Eppendorf tubes. The tubes were briefly centrifuged to place the sample towards the bottoms or sides of the tube. RNA extraction and DNase treatment was performed using the RNeasy FFPE kit (Qiagen) according to the manufacturer's instructions, using the Xylene protocol for the paraffin removal step and 30 µL RNase-free H2O for the elution step. RNA was initially quantified by NanoDrop. Subsequent total RNA quantification and quality assessment was performed by Bioanalyzer 2100 (Agilent Technologies). Samples were excluded from the study if >20% of the RNA fragments were >200 base pairs in length, as determined with the Bioanalyzer software. Library preparation was performed by the Genome Technology Access Center (GTAC) at Washington University. Library preparation used 150 ng of total RNA. ds-cDNA was generated using the Ovation RNA-seq FFPE system (Nugen, per manufacturer's instructions). cDNA was blunt ended, had an A base added to the 3' ends, and then had Illumina sequencing adapters ligated to the ends. Ligated fragments were then amplified for 12 cycles using primers incorporating unique index tags.
attributes.tag: ENA first public
attributes.value: 2018-01-01
attributes.tag: ENA last update
attributes.value: 2017-05-26
attributes.tag: External Id
attributes.value: SAMEA104101968
attributes.tag: INSDC center alias
attributes.value: Washington University School of Medicine
attributes.tag: INSDC center name
attributes.value: Washington University School of Medicine
attributes.tag: INSDC first public
attributes.value: 2018-01-01T17:01:40Z
attributes.tag: INSDC last update
attributes.value: 2017-05-26T09:34:24Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-5783:WUCA-Cont-29
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: common name
attributes.value: human
attributes.tag: disease
attributes.value: normal
attributes.tag: organism part
attributes.value: small intestine
attributes.tag: sample name
attributes.value: E-MTAB-5783:WUCA-Cont-29
attributes.tag: sampling site
attributes.value: terminal ileum
attributes.tag: specimen with known storage state
attributes.value: paraffin specimen