An improved in-vitro regeneration protocol using scutellum of mature and immature embryos of wheat

Abstract

Generally, conventional practices take a long time to improve agronomic traits in common wheat (Triticum aestivum L.). However, biotechnological tools provide the best opportunity to enhance crop potential within a limited time. A simple and repeatable protocol for wheat regeneration is the priority for transforming the genetic makeup of elite wheat cultivars. Among 10 types of callus induction media (CIM), CIM4 with supplemented Murashige and Skoog (MS) media+ dicamba (2 mg/l) has 98.22 and 97.33% callus induction efficiency (CIE) in immature and mature scutellum, respectively. Precocious germination, considered a key barrier during callus induction, was fully controlled by excising the embryonic axis. Among 29 different shoot induction media (SIM) assessed in the study, SIM 29 containing zeatin 5 mg/l, 6-benzylaminopurine (BAP) 5 mg/l, 2,4-dichlorophenoxyacetic acid (2,4-D) 0.25 mg/l,1-naphthaleneacetic acid (NAA) 0.25 mg/l, and copper sulfate (CuSO4) 20 mg/l was found most responsive for shoot induction. The combined effect of cytokinin and CuSO4 resulted in an improved wheat regeneration response in two popular Indian wheat varieties, namely HD2967 and HD3171. Rooting was observed at 100% using IAA and IBA @ 0.2 mg/l concentration in root induction media (RIM). The whole in-vitro regeneration protocol from embryo to transfer in the greenhouse took 8 to 9 weeks. The in-vitro regeneration protocol developed in the present study could further be utilized to improve the genetic makeup of elite wheat cultivars utilizing genetic engineering tools.