A new generation of human artificial chromosomes for functional genomics and gene therapy. (49/58)

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LINE1 family member is negative regulator of HLA-G expression. (50/58)

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Centromere architecture breakdown induced by the viral E3 ubiquitin ligase ICP0 protein of herpes simplex virus type 1. (51/58)

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Telomere length homeostasis and telomere position effect on a linear human artificial chromosome are dictated by the genetic background. (52/58)

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Trans-chromosomic mice containing a human CYP3A cluster for prediction of xenobiotic metabolism in humans. (53/58)

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Comparative study of artificial chromosome centromeres in human and murine cells. (54/58)

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Organization of synthetic alphoid DNA array in human artificial chromosome (HAC) with a conditional centromere. (55/58)

Human artificial chromosomes (HACs) represent a novel promising episomal system for functional genomics, gene therapy, and synthetic biology. HACs are engineered from natural and synthetic alphoid DNA arrays upon transfection into human cells. The use of HACs for gene expression studies requires the knowledge of their structural organization. However, none of the de novo HACs constructed so far has been physically mapped in detail. Recently we constructed a synthetic alphoid(tetO)-HAC that was successfully used for expression of full-length genes to correct genetic deficiencies in human cells. The HAC can be easily eliminated from cell populations by inactivation of its conditional kinetochore. This unique feature provides a control for phenotypic changes attributed to expression of HAC-encoded genes. This work describes organization of a megabase-size synthetic alphoid DNA array in the alphoid(tetO)-HAC that has been formed from a ~50 kb synthetic alphoid(tetO)-construct. Our analysis showed that this array represents a 1.1 Mb continuous sequence assembled from multiple copies of input DNA, a significant part of which was rearranged before assembling. The tandem and inverted alphoid DNA repeats in the HAC range in size from 25 to 150 kb. In addition, we demonstrated that the structure and functional domains of the HAC remains unchanged after several rounds of its transfer into different host cells. The knowledge of the alphoid(tetO)-HAC structure provides a tool to control HAC integrity during different manipulations. Our results also shed light on a mechanism for de novo HAC formation in human cells.  (+info)

Re-engineering an alphoid(tetO)-HAC-based vector to enable high-throughput analyses of gene function. (56/58)

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