Elucidating molecular interaction leading to successful grafting of in vitro regenerated shoots of three chickpea (Cicer arietinum L.) cultivars

Chickpea is a crucial pulse crop worldwide, serving as a significant source of protein. It is predominantly cultivated in the Indian subcontinent, where diverse consumption patterns have driven increased production demand. An efficient regeneration protocol is required for genetic manipulation of chickpea. We have improvised the regeneration protocol of three chickpea cultivars viz. JG62, BG212 and C-104. Twelve media combinations for shoot induction were utilized to regenerate shoots. These regenerated shoots were successfully grafted onto the rootstocks of their respective cultivars, achieving a grafting success rate of 60–65%. Highest grafting efficiency was observed with 10-day old rootstocks and at a height of 4 cm above the base. To get insight into the grafting process, we performed High-performance liquid chromatography (HPLC) for auxin hormone estimation and expression studies involving a set of genes from auxin pathway including efflux transporters like CaPIN1 (for tissue polarity), CaPIN3 (exhibiting gravity-sensing in tissues), CaHB8 (procambial activity marker), CaTIR1 (transport inhibitor response1), CaARF12 and CaARF14 (auxin-responsive factor), CaH4 (cell division marker) and CaALF4 (aberrant lateral root formation 4) and also CaWIND1 (wound-induced dedifferentiation1) from cytokinin hormone pathway. Semi-quantitative and real time PCR showed relative abundance of these transcripts at graft union site revealing involvement of auxin pathway. An increase in endogenous auxin was also observed at 5 DAG. Thus, our finding supports the canalization hypothesis for successful graft union by regeneration of continuous vascular cambium wherein, high grafting efficiency is achieved by polar auxin transport that regulates vascular reconnection.