1. An optimized method for high-titer lentivirus preparations without ultracentrifugation.
- Author
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Jiang, Wei, Hua, Rui, Wei, Mengping, Li, Chenhong, Qiu, Zilong, Yang, Xiaofei, and Zhang, Chen
- Subjects
ULTRACENTRIFUGATION ,HIV infection complications ,NUCLEOTIDE sequence ,DNA analysis ,LENTIVIRUS diseases ,DIAGNOSIS - Abstract
Lentiviral technology has proven to be a powerful tool to express exogenous genes in dividing and non-dividing cells. Currently, most protocols for generating high-titer lentivirus require ultracentrifugation, which can be an instrumental barrier for routine operations in a laboratory. In this study, the effect of relative centrifugal force (RCF) on the concentration efficiency of the lentivirus was systematically explored, and it was found that sucrose gradient centrifugation with a relatively low speed (≤10,000 g) robustly produces a high-titer virus (up to 2 × 10
8 TU/ml). The optimal sucrose concentration is 10%, and the recovery rate of the functional virus is greater than 80%. The infection efficiency of both concentrated and un-concentrated lentivirus decreases rapidly when the viruses are stored at 4 °C (τ ≈ 1.3 days) or subjected to multiple freeze-thaw cycles (τ = 1.1 rounds). In summary, we describe an efficient and easy-to-handle protocol for high-titer lentivirus purification. [ABSTRACT FROM AUTHOR]- Published
- 2015
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