A Novel Direct PCR Lysis Buffer Can Improve PCR from Meat Matrices
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A Novel Direct PCR Lysis Buffer Can Improve PCR from Meat Matrices Feng Guan 1 & Yuting Jin 1 & Jin Zhao 1 & Juntao Ai 2 & Yuanyuan Luo 1 Received: 28 May 2018 / Accepted: 9 August 2018 # Springer Science+Business Media, LLC, part of Springer Nature 2018
Abstract Molecular technologies based on PCR have been widely used in many biological analysis fields, especially for genetic analysis and DNA barcoding. In this study, a rapid DNA lysis liquid was formulated without any purification step from fresh and processed meat, suitable for conventional PCR amplification. Three different lysis liquid formulas were designed for selection and further optimization for direct PCR and absorbance spectra; DNA concentration and performance in PCR were used to assess the effect of each formula. The results indicated that the formula containing NaOH, EDTA, SDS, Tween 20, and Tris-HCl achieved the best results, and the optimized formula met the need of practical PCR applications. The protocol provided a rapid lysis buffer for DNA replication from any meat samples. The performance of the final formula resulted in high DNA lysis efficiencies for all the tested meat samples and the PCR amplification efficiencies were similar to isolated DNA template using a commercial kit. The whole process can be completed in 30 min. Therefore, this study provides a simple, alternative, cost-effective fast solution for meat molecular analysis based on DNA. Keywords Meat samples . Lysis liquid . Direct PCR . DNA amplification
Introduction PCR amplifications have been widely used in almost all molecular biological techniques because it is reliable, robust, and rapid, especially for DNA-based analysis. Numerous changes have been implemented for improving the efficiency of PCR analysis ever since the first protocol to extract and amplify DNA was developed. These changes have revolutionized the application of PCR in molecular biology field. One of the notable applications is DNA barcoding performed for species identification (Kumar et al. 2015; Rahmati et al. 2016): both for animals (Nagalakshmi et al. 2016; Rahmati et al. 2016; Safdar and Junejo 2016) and plant species (Madesis et al. 2014). In general, the premise of most of the molecular technologies based on PCR is to extract DNA from many kind of matrices, such as animal tissues (Fernandes et al. 2017; Harikai and Shinomiya 2014; Kitpipit et al. 2014b; Lockley and Bardsley 2000; Madesis et al. 2014; Rahmati et al. 2016; Song et al. 2017; Toubanaki and Karagouni 2017; Yan et al.
* Feng Guan [email protected] 1
College of Life Sciences, China Jiliang University, Hangzhou 310018, China
2
Beijing Vocational College of Agriculture, Beijing 10000, China
2016), bone (Yancy et al. 2005), plant tissues, and food made of them (Madesis et al. 2014). However, the methods employed to extract DNA are complicated and require timeconsuming purification process. In addition, several commercial kits have been developed for different materials and highthroughput DNA extraction, and these kits have good results
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