Skip to main content
. 2016 Jul 25;236:178–183. doi: 10.1016/j.jviromet.2016.07.022

Fig 2.

Fig 2

(a,b) Transformed S. cerevisiae colonies were screened for the MERS-CoV/pYES1L plasmid vector by PCR (Primer Set 1: F- GTGCTGATGACGAAGGCTTCATCACATTAAAG-AACAATCTATA and R- AGTTGCAAACCTTATACGGTATGGTTGCAACTTTCTTAAAGGAC, Primer Set 2: F- AAGCTTGCACCATGTTGAAACTTTTGGTGGAAGTGCGCTTGT and R- TTAACTGCGGCGAGGCAGCATATGGCATATGTTGCCAAACTCTAAACCAAATAC). Each well (1–23) represents a single colony; positive colonies were defined as those producing an amplicon for both Primer set 1 and Primer set 2. (c) Cellular supernatant was withdrawn from cells infected with either rescued rMERS-CoV virus or rescued rSARS-CoV virus and used in a RT-PCR assay to detect a region of the MERS-CoV polymerase gene (Primer set: F-CAAGATGAACTTTTTGCCATGACAAAGCGTAACGTCATTCC and R- TGTGAGAGGGTAAGCATCTATAGCCAAAGACACAAACCGCT). The MERS-CoV polymerase gene was detected in Lane 1 (rMERS-CoV supernatant) and not Lane 2 (rSARS-CoV supernatant). Lane 3 serves as a positive control, it shows PCR amplification of the MERS-CoV polymerase gene from the infectious clone system. (d) The polymerase gene fragment amplified from rMERS-CoV infected cell supernatant was sequenced and used in a phylogenetic comparison (ClustalW Algorithim) with six coronaviruses: CoV-HKU4-1 (NC_009019.1), CoV-HKU5-1 (NC_009020.1), SARS-CoV/Tor 2 (NC_004718.3), CoV-Neoromicia/PML (KC_869678.4), CoV-229E (NC_002645.1) and MERS-CoV/EMC/2012 (NC_019843.3).