PMC:1160574 / 6678-8053 JSONTXT

{"target":"http://pubannotation.org/docs/sourcedb/PMC/sourceid/1160574","sourcedb":"PMC","sourceid":"1160574","source_url":"http://www.ncbi.nlm.nih.gov/pmc/1160574","text":"Gene-Expression Analyses\n\nRT-PCR analysis\nTotal RNA was extracted by using the RNeasy kit and DNase I treatment (Qiagen, Valencia, California, United States). Total RNA (2 μg each) was reverse transcribed (SuperScript; Invitrogen, Carlsbad, California, United States). PCR conditions were optimized and linear amplification range was determined for each primer by varying annealing temperature and cycle number. PCR products were identified by size, and identity was confirmed by DNA sequencing. Primer sequences, cycle numbers, and annealing temperatures are provided in Table S1.\n\nAffymetrix analysis\nTotal RNA (5 μg) from primary MSCs, from hESMPC-H9.1, hESMPC-H1.2, and three samples of undifferentiated hESCs (H1; passages 42–46), were processed by the Memorial Sloan-Kettering Cancer Center Genomics Core Facility and hybridized on Affymetrix (Santa Clara, California, United States) U133A human oligonucleotide arrays. Data were analyzed using MAS5.0 (Affymetrix) software. Transcripts selectively expressed in each of the mesenchymal cell populations (MSC, hESMPC-H9.1, and hESMPC-H1.2) were defined as those called “increased” by the MAS5.0 algorithm in each of three comparisons with independent samples of undifferentiated hESCs. A Venn diagram was generated to visualize overlap in gene expression. Further statistical analyses were performed as described below.\n","divisions":[{"label":"Title","span":{"begin":0,"end":24}},{"label":"Section","span":{"begin":26,"end":581}},{"label":"Title","span":{"begin":26,"end":41}},{"label":"Section","span":{"begin":583,"end":1374}},{"label":"Title","span":{"begin":583,"end":602}}],"tracks":[{"project":"CellFinder","denotations":[{"id":"T175","span":{"begin":1082,"end":1093},"obj":"CellLine"},{"id":"T176","span":{"begin":633,"end":637},"obj":"CellType"},{"id":"T177","span":{"begin":1072,"end":1076},"obj":"CellLine"},{"id":"T178","span":{"begin":708,"end":713},"obj":"CellType"},{"id":"T179","span":{"begin":657,"end":668},"obj":"CellLine"},{"id":"T180","span":{"begin":651,"end":655},"obj":"CellLine"},{"id":"T181","span":{"begin":664,"end":668},"obj":"CellLine"},{"id":"T182","span":{"begin":1089,"end":1093},"obj":"CellLine"},{"id":"T183","span":{"begin":1234,"end":1239},"obj":"CellType"},{"id":"T184","span":{"begin":896,"end":901},"obj":"Species"},{"id":"T185","span":{"begin":715,"end":717},"obj":"CellLine"},{"id":"T186","span":{"begin":644,"end":655},"obj":"CellLine"},{"id":"T187","span":{"begin":1030,"end":1041},"obj":"Anatomy"},{"id":"T188","span":{"begin":1065,"end":1076},"obj":"CellLine"}],"attributes":[{"subj":"T175","pred":"source","obj":"CellFinder"},{"subj":"T176","pred":"source","obj":"CellFinder"},{"subj":"T177","pred":"source","obj":"CellFinder"},{"subj":"T178","pred":"source","obj":"CellFinder"},{"subj":"T179","pred":"source","obj":"CellFinder"},{"subj":"T180","pred":"source","obj":"CellFinder"},{"subj":"T181","pred":"source","obj":"CellFinder"},{"subj":"T182","pred":"source","obj":"CellFinder"},{"subj":"T183","pred":"source","obj":"CellFinder"},{"subj":"T184","pred":"source","obj":"CellFinder"},{"subj":"T185","pred":"source","obj":"CellFinder"},{"subj":"T186","pred":"source","obj":"CellFinder"},{"subj":"T187","pred":"source","obj":"CellFinder"},{"subj":"T188","pred":"source","obj":"CellFinder"}]}],"config":{"attribute types":[{"pred":"source","value type":"selection","values":[{"id":"CellFinder","color":"#9eec93","default":true}]}]}}