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DNASTAR- DNA Cloning Software - Sequence Analysis …
The identification of restriction enzymes capable ofrecognizing and cleaving DNA at specific sites has been acornerstone of modern biotechnology (). Genomic DNA digested by restrictionenzymes becomes DNA fragments of varied lengths, creating apersonalized signature called restriction fragment lengthpolymorphisms (RFLPs). Prior to the widespread use ofhigh-throughput sequencing and genotyping methods, RFLP was one ofthe major assays for pinpointing genomic regions responsible forvarious phenotypic traits (). Thistechnology has led toward the discovery of the gene,the first disease-bearing gene ever identified by positionalcloning (,). The method has also been used invarious clinical assays, including the diagnosis of sickle cellanemia ().
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AB - Stem cells that can give rise to neurons, astroglia, and oligodendroglia have been found in the developing and adult central nervous system (CNS) of rodents. Yet, their existence within the human brain has not been documented, and the isolation and characterization of multipotent embryonic human neural stem cells have proven difficult to accomplish. We show that the developing human CNS embodies multipotent precursors that differ from their murine counterpart in that they require simultaneous, synergistic stimulation by both epidermal and fibroblast growth factor-2 to exhibit critical stem cell characteristics. Clonal analysis demonstrates that human C NS stem cells are multipotent and differentiate spontaneously into neurons, astrocytes, and oligodendrocytes when growth factors are removed. Subcloning and population analysis show their extensive self-renewal capacity and functional stability, their ability to maintain a steady growth profile, their multipotency, and a constant potential for neuronal differentiation for more than 2 years. The neurons generated by human stem cells over this period of time are electrophysiologically active. These cells are also cryopreservable. Finally, we demonstrate that the neuronal and glial progeny of long-term cultured human CNS stem cells can effectively survive transplantation into the lesioned striatum of adult rats. Tumor formation is not observed, even in immunodeficient hosts. Hence, as a consequence of their inherent biology, human CNS stem cells can establish stable, transplantable cell lines by epigenetic stimulation. These lines represent a renewable source of neurons and glia and may significantly facilitate research on human neurogenesis and the development of clinical neural transplantation.