In vitro plant regeneration from cotyledonary nodes of recombinant inbred lines of lentil

An efficient and reproducible in vitro regeneration protocol for lentil was developed. For shoot regeneration, cotyledonary node explants of ten elite genotypes were cultured in an inverted orientation on different shoot regeneration media that consisted of Murashige and Skoog (MS) medium supplemented with 1 mg L−1 6-benzylaminopurine (BAP) (M1), 1 mg L−1 BAP + 0.45 mg L−1 indole-3-acetic acid (IAA) (M2), and 2 mg L−1 BAP (M3). High percentages of shoot regeneration ranging from 80 to 100% on M1 and M3 media and from 50 to 100% on M2 medium were induced. M1 was the most efficient shoot regeneration medium for most genotypes tested. For rooting, in vitro and in vitro–in vivo methods were used. Low and variable rooting percentages ranging from 0 to 45% were recorded with in vitro–in vivo method. Efficiency of rooting on in vitro medium varied depending on the medium in which shoots had been previously regenerated and the genotype tested. When M1 medium was used, high rooting percentages (over 40%) for most genotypes except for microsperma genotypes were found. When the 10 genotypes were screened for good regeneration performance using M1 medium, 2 main clusters and 3 subgroups within one of the clusters were formed based on similarities respect of the number of regenerated shoots per explant and rooting percentages. Subgroup 1 composed by A1146 genotype produced the highest number of shoots per explant (6.17 shoots) and a high rooting percentage (60%) so was selected for further transformation and use as a potential commercial variety.

► An efficient and reproducible in vitro regeneration protocol for lentil was developed.
► M1 containing 1 mg L−1 BAP was the most efficient shoot regeneration medium for most genotypes tested.
► We reached high rooting percentages using a growth-regulator-free medium (M0) for rooting.
► Rooting percentages were lower for microsperma types.