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. 2017 Dec 19;8:2137. doi: 10.3389/fpls.2017.02137

Figure 6.

Figure 6

Transcription start site and NbKPILP mRNA 5'-UTR identification. (A) Schematic representation of the 5′-RACE procedure: cDNA preparation using addition of non-templated dCMP residues to the 3′-end of full-length cDNA by reverse transcriptase (Schmidt and Mueller, 1999) followed by the cDNA second strand synthesis using “PlugOligo” primer and further amplification using “M1” and sequence-specific primers. The position of the primers is marked with arrows. AUG and UAA correspond to start and stop codons of the NbKPILP ORF. Sequence complementary to PlugOligo primer designated with gray box; double line with an arrowhead stands for the forward M1 primer; pr1 and pr2, positions of the reverse primers for PCR. (B) Agarose gel electrophoresis of PCR products obtained using the adapter primer and pr1 or pr2. Lanes correspond to the following samples: 1—leaves from the plant incubated in darkness for 96 h, 2—leaves from the plant incubated at a normal photoperiod for 24 h after 96-h darkness, 3—leaves 3 days after agroinjection with 35S-NbKPILP. (C) The nucleotide sequence of the NbKPILP mRNA 5′-UTR. The putative uORF is underlined. TSS, transcription start site.