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Construction and Detection of a Novel Type of Recombinant Human rAAV2/2-ND4

   
@article{YKXB3734,
	author = {Han Pei 和 Xing Wan 和 Weikun Hu 和 Xiaoyan Dong 和 Bin Li},
	title = {Construction and Detection of a Novel Type of Recombinant Human rAAV2/2-ND4},
	journal = {眼科学报},
	volume = {28},
	number = {2},
	year = {2016},
	keywords = {},
	abstract = {Purpose:To construct a novel AAV-mediated gene delivery of the human ND4 complex I subunit and to detect its expression level in mitochondria for potential application in gene therapy for Leber’s hereditary optic neuropathy (LHON). Methods:A novel type of normal human ND4 gene was synthesized artificially to contain a mitochondrial targeting sequence that induces the translocation of this gene into mitochondria.This recombinant adeno-associated virus type 2/serotype 2 (rAAV2/2)-mediated NADH dehydrogenase subunit 4 (ND4) gene was constructed, purified, condensed, and amplified by PCR.The physical titer of rAAV2/2-ND4 was determined by slot-blot hybridization using a digoxigeninlabeled H1 probe.Expression of ND4 in mitochondria was evaluated by immunofluorescence. Results: The constructed rAAV2/2-ND4 specifically amplified the target gene band of ND4 and the physical titer of ND4 gene was 1.0×1011 vg/mL,confirming that the recombinant adenovirus vector contained the ND4 target gene. Expression of the ND4 gene was detected in mitochondria by immunofluorescence. Conclusion:A new type of rAAV2/2-ND4 was successfully constructed and may have potential in gene therapy for LHON.},
	url = {https://ykxb.amegroups.com/article/view/3734}
}