Generation of miR156 knockdown lines and expression analysis of putative miRNAs involved in potato tuberisation

Abstract

Micro RNAs (miRNAs) are small, non-coding RNAs that regulate different developmental pathways in plants including tuberisation in potato. Previous reports have demonstrated the role of miR156 and miR172 in regulating tuberisation pathway in potato, where miR156 and miR172 facilitate tuber formation under tuber inductive Short Day (SD) conditions. Functional studies of these miRNAs in potato were carried out earlier by employing over expression strategies. To gain more insights in miR156 function in potato development, we have attempted miR156 knockdown approach. In this regard, we have cloned and generated mimicry-miR156 (MIM156) lines in potato and tobacco. MIM-miR156 transgenic plants were successfully generated and were confirmed by RT-qPCR. Currently, we are analyzing and comparing the miR156 knockdown phenotype of potato and tobacco transgenics. Apart from the reports of miR156 and miR172 in potato, many reports have demonstrated the presence of numerous miRNAs in potato by employing bioinformatic and deep sequencing approaches. However, their function in potato tuberisation pathway is not known. Based on the previous reports, we have shortlisted few miRNAs with an aim to identify other miRNAs possibly involved in tuberisation pathway. Tuberization being a photoperiod dependent phenomenon, we have carried out SD-LD photoperiod dependent expression pattern of these shortlisted miRNAs, where miR126, miR394, miR171, miR157 and miR159 showed differential expression. These miRNAs could potentially be involved in tuberisation pathway awaiting future investigations. In this report, we aim to understand the miRNA mediated regulation of tuberisation pathway by (i) understanding the role of miR156 using MIM156 transgenic approach and (ii) SD-LD dependent expression pattern of different miRNAs in potato. Overall, this work will help to identify the role of other miRNA candidates potentially involved in tuberisation pathway.