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. 2018 Jun 29;9:833. doi: 10.3389/fpls.2018.00833

FIGURE 2.

FIGURE 2

Cis/trans test of heat-induced ZmbZIP60 expression. (A) F1 hybrid lines were obtained from crosses of B73 × high expressing RILs with CML52 type promoters. ZmbZIP60 expression was heat induced and the RT-PCR product was sequenced to determine the relative expression of the two alleles. (B) DNA sequence analysis of the B73 type and the CML52 type alleles. Six SNPs in the 5′-UTR were used to distinguish the contribution of the two different alleles in the RNA transcript population. Green highlights the coding region and purple highlights sequence variation between the alleles. Boxes indicate the location of the primers used in the RT-PCR analysis. (C) Sequence profile of RT-PCR products from the analysis of hybrid line 1 derived from a cross of B73 × Z008E0012. RNA extracted from seedlings incubated at 26°C served as a control, while the experimental sample was extracted from seedlings incubated for 1 h at 39°C. The area under the peaks for the SNPs (pointed out by arrows) was measured and the ratio of the two different alleles was calculated. (D) RT-PCR analysis of the RNA extracted from control hybrid seedlings incubated at 26°C and those subjected to heat treatment (39°C, 1 h). Bar graph represents the normalized contribution of the two alleles (CML type and B73 type) to the RNA population in the control and heat-treated seedlings.