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. 2012 Dec 26;7(12):e52734. doi: 10.1371/journal.pone.0052734

Figure 4. Schematic diagram of the construction of reporters in silencing assay.

Figure 4

The upper panel shows the construction of URA5-miRs or URA5-miR-ms. Two pair of primers, URA5-XhoI-S/URA5-miRs-BamHI and URA5-BamHI-S/URA5-XbaI-A, were used to PCR amplify the ORF and the terminator regions of URA5, respectively. Among the primers, URA5-miRs-BamHI harbored the sequence of miRs or miR-ms through primer design. The two PCR fragments were then ligated after digested by BamH I. Similarly, the construction of CLC-miR1/2 and CLC-miR1-m/miR2-m was made (the bottom part). The position of miRs or miR-ms is indicated by the boxes. Restriction sites are in small letters. Arrows mark the position and direction of the primers. Start codons and stop codons of URA5 and CLC1 are underlined. For detailed description, see the section of Materials and Methods.