Abstract
REVERSE genetic approach was used to isolate and characterize Medicago truncatula containing "knockout" mutations in gene involved in nodulation process and in many other physiological processes. More than 60 Tnt1-Flanking sequence tags (FSTs) ranging from similar to 70bp to similar to 600bp were isolated, sequenced and submitted to Genebank referring for akt1 mutant characterization. The proper Tnt1-insertion was mapped in chromosome number 8 of Medicago truncatula genome. It was precisely identified upstream the base number 141 and downstream of the ATG start codon of Medicago truncatula potassium channel AKT1 gene (MtAKT1). MtAKT1 gene encodes inwardly rectifying potassium channel was isolated, sequenced, and submitted to Genebank with accession number MN649185.1.M. truncatula akt1 mutant is achieving higher number of deformed root nodules and shorter root length compared to wild type. akt1 nodules exhibited reduced size occupied with un-differentiated cells and abnormalities in symbiotic nodule zones. Non-functional nitrogen fixation zone and compacted infection zone were observed as well. AKT1 null mutant exhibits attenuation in its ability to maintain the proper K+ and Na+ ions content in akt1 seedlings which is 2-3 fold less than wild type seedlings,. In contrast, akt1 seedlings showed three-fold increase in Ca++ ion concentration compared to wild type.
In conclusion Medicago truncatula mutant, akt1 is Tnt1-retrotransposon mutant impaired in inwardly rectifying potassium channel AKT1. As the first reported AKT1 null mutant was isolated from legume plants, this mutant is displaying abnormalities in nitrogen fixation organ and affecting ions uptake.