Microarray-based comparative genomic hybridization (array CGH) is a high-resolution and comprehensive method for detecting both genome-wide and chromosome-specific copy-number imbalance. We have developed an array CGH analysis system (consisting of an array CGH chip plus its exclusive analysis software) for constitutional genetic diagnosis and have evaluated the suitability of our system for molecular diagnosis using preand postnatal clinical samples. In a blind study, each of the 264 sample karyotypes identified by array CGH analysis was consistent with that identified by traditional karyotype analysis -with one exception, case (47, XXX) -and we were able to identify origins, such as small supernumerary marker chromosomes, which cannot be determined by conventional cytogenetics. We also acquired very accurate, fast and reliable results using a diminutive amount of clinical samples. Taken together, the array CGH platform developed in this study is a rapid, powerful and sensitive technology for pre-and postnatal diagnosis using a very small amount of clinical sample.
To obtain an enhanced population of cardiomyocytes from differentiating mouse embryonic stem (ES) cells, we confirmed the role of noggin treatment during the cardiac differentiation of mouse ES cells. ES cells were cultured in ES medium containing both noggin and LIF for 3 days on the mouse embryonic fibroblast feeder layer, followed by dissociated and suspension culture without LIF to form the embryoid body (EB). The next day, noggin was eliminated and EBs were cultured continuously. Noggin treated ES cells showed a relatively rapid increase of cardiac marker genes, while the vehicle (PBS) treated group showed no significant cardiac marker expression at 4 days after the EB formation. Furthermore, Noggin treated ES cells showed 68.00±9.16% spontaneous beating EBs at 12 days after the EB formation. To develop a more efficient cardiomyocyte differentiation method, we tested several known cardiogenic reagents (ascorbic acid, 5’-Azacytidine, LiCl, oxytocin, FGF2 and PDGF-BB) after noggin induction or we cultured noggin treated ES cells on various extracellular matrixes (collagen, fibronectin and Matrigel). Quantitative RT-PCR and immunocytochemistry results showed a significantly increased cardiac differentiation rate in the FGF2 treated group. Differentiation on the collagen extracellular matrix (ECM) could slightly increase the cardiac differentiation efficiency. These results show the possibilities for the establishment of selective differentiation conditions for the cardiac differentiation of mouse ES cells.
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