A low-cost and open-source protocol to produce key enzymes for molecular detection assays. Mendoza-Rojas et al. (2021)

Here, we describe a detailed step-by-step protocol for the expression, purification, quantification, and activity determination of key enzymes for molecular detection of pathogens. Based on previous reports, we optimized the protocol for LbCas12a, Taq DNA polymerase, M-MLV reverse transcriptase, and TEV protease to make it compatible with minimal laboratory equipment, broadly available in low- and middle-income countries. The enzymes produced with this protocol have been successfully used for molecular detection applications.
For complete details on the use of this protocol, please refer to Alcantara et al. (2021).

本研究详细描述了一套分步标准化实验流程，用于病原体分子检测关键酶的表达、纯化、定量与活性测定。基于既往研究报道，我们针对LbCas12a、Taq DNA聚合酶（Taq DNA polymerase）、M-MLV反转录酶（M-MLV reverse transcriptase）以及TEV蛋白酶（TEV protease）优化了该方案，使其可适配低收入和中等收入国家广泛可得的简易实验室设备。采用本流程制备的酶已成功应用于分子检测相关实验。
如需获取本方案的完整使用细节，请参阅Alcantara等人（2021）的研究。