![]() ![]() ![]() The most frequently used is plasmid transfection, where 2 to 3 different plasmids and DNA sequences are transfected into the cell. The ratios of these different viral and cellular helper proteins, as well as the AAV2 replication protein, help to dictate the overall number of particles packaged, the number of particles that contain DNA, and often the integrity or completeness of the genome that’s packaged within these capsids.Ĭurrently, multiple methods are utilized to deliver each of these helper components and replication and capsid sequences into the cell. The replicated DNA sequences are then packaged into the AAV capsid and harvested from the cells and/or supernatant through the purification process. These proteins are critical in AAV production, assisting in multiple functions including limiting replication of the packaging cell, expressing viral capsid proteins, and increasing production of the Cis DNA sequences. When these helper virus sequences are stressed, they trigger expression of both cellular factors critical in AAV production and activate the four different AAV Rep proteins (48, 52, 68, 78). Random shuffling of capsid sequences to generate new novel capsids andĪAV vectors are produced through the introduction of helper virus and AAV replication components into the production cell line.Directed evolution, which incorporates components of multiple AAV serotypes into the capsid.Capsid proteins have been further modified to increase transduction, targeting specificity, and efficacy in vivo by methods that include: To date, multiple AAV serotypes targeting different organs including brain, eye, lung liver, skeletal muscle, and heart have been discovered and characterized. The utilization of AAV as a gene therapy vector has increased due to its relatively limited immunotoxicity and wide range of tissue tropism. Introduction: challenges in improving AAV productivity & scalability ![]()
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