PCR Amplicon Sequencing and Purification

The PCR product was purified using the ExoSAP clean-up kit (Thermofisher Scientific, USA). The PCR amplicons were then sequenced using the BigDye Cycle Sequencing Kit (Applied Biosystems, USA). The sequencing was performed with the same forward primers as were used for the RT-PCR [43 (link)]. After the sequencing reaction, an ethanol precipitation was performed and the final product was loaded in ABI PRISM 3130 automated sequencer (Applied Biosystems, USA) [44 (link)].

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Sadiq A., Bostan N., Bokhari H., Matthijnssens J., Yinda K.C., Raza S, & Nawaz T. (2019). Molecular characterization of human group A rotavirus genotypes circulating in Rawalpindi, Islamabad, Pakistan during 2015-2016. PLoS ONE, 14(7), e0220387.

Publication 2019

ExoSAP clean-up kit BigDye Cycle Sequencing Kit ABI PRISM 3130

Ethanol Primers Prism Rt pcr

Corresponding Organization : KU Leuven

Top 5 similar protocols

Variable analysis

independent variables

Forward primers used for RT-PCR

dependent variables

DNA sequence of PCR amplicons

control variables

Ethanol precipitation step
Use of ABI PRISM 3130 automated sequencer

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