Vadim Timerbaev1,2,3, Sergey Dolgov4,5,6. 1. Branch of the Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Pushchino, 142290, Russia. timerbaev@gmail.com. 2. Nikita Botanical Gardens-National Scientific Center, Russian Academy of Sciences, Yalta, 298648, Russia. timerbaev@gmail.com. 3. All-Russia Research Institute of Agricultural Biotechnology, Russian Academy of Sciences, Moscow, 127550, Russia. timerbaev@gmail.com. 4. Branch of the Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, Pushchino, 142290, Russia. 5. Nikita Botanical Gardens-National Scientific Center, Russian Academy of Sciences, Yalta, 298648, Russia. 6. All-Russia Research Institute of Agricultural Biotechnology, Russian Academy of Sciences, Moscow, 127550, Russia.
Abstract
MAIN CONCLUSION: The tomato ELIP gene promoter is mainly active in the ripening fruit. Considering its high activity, the promoter could be used for molecular breeding of plants in the future. The ability to obtain new varieties of transgenic plants with economically valuable traits relies on a high level of target gene expression, which is largely controlled by a gene promoter. Hence, research aimed at finding and characterizing new tissue-specific promoters that direct gene expression in specific plant tissues or at certain developmental stages has become the most important field of plant biotechnology. Here, we cloned and characterized the promoter of the early light-inducible protein (ELIP) gene from tomato (Solanum lycopersicum cv. Yalf). ELIPs are produced in the presence of light and putatively function in the chloroplast-to-chromoplast conversion, playing a photorepairing role in the photosynthetic system. Analysis of the promoter sequence revealed multiple cis-acting elements related to light responsiveness, and other motifs involved in plant hormone response and circadian control. To determine the functionality of the promoter, seven 5'-deletion variants were fused with the β-glucuronidase (GUS) reporter gene and introduced into tomato. Histochemical analysis of transgenic tomato plants revealed different levels of GUS activity in most analyzed tissues, depending on the promoter fragment used. The intensity of staining was considerably higher in ripening fruits than in unripe and non-fruit tissues. Quantitative analysis indicated that the level of GUS activity with the longest (full-length) version of the ELIP promoter in ripened fruits was comparable to that in plants expressing the constitutive CaMV35S promoter. Further, the location of both negative and positive regulatory motifs was identified. The described ELIP promoter is a potential tool for various applications in plant biotechnology.
MAIN CONCLUSION: The tomato ELIP gene promoter is mainly active in the ripening fruit. Considering its high activity, the promoter could be used for molecular breeding of plants in the future. The ability to obtain new varieties of transgenic plants with economically valuable traits relies on a high level of target gene expression, which is largely controlled by a gene promoter. Hence, research aimed at finding and characterizing new tissue-specific promoters that direct gene expression in specific plant tissues or at certain developmental stages has become the most important field of plant biotechnology. Here, we cloned and characterized the promoter of the early light-inducible protein (ELIP) gene from tomato (Solanum lycopersicum cv. Yalf). ELIPs are produced in the presence of light and putatively function in the chloroplast-to-chromoplast conversion, playing a photorepairing role in the photosynthetic system. Analysis of the promoter sequence revealed multiple cis-acting elements related to light responsiveness, and other motifs involved in plant hormone response and circadian control. To determine the functionality of the promoter, seven 5'-deletion variants were fused with the β-glucuronidase (GUS) reporter gene and introduced into tomato. Histochemical analysis of transgenic tomato plants revealed different levels of GUS activity in most analyzed tissues, depending on the promoter fragment used. The intensity of staining was considerably higher in ripening fruits than in unripe and non-fruit tissues. Quantitative analysis indicated that the level of GUS activity with the longest (full-length) version of the ELIP promoter in ripened fruits was comparable to that in plants expressing the constitutive CaMV35S promoter. Further, the location of both negative and positive regulatory motifs was identified. The described ELIP promoter is a potential tool for various applications in plant biotechnology.