Exogenous Polyamines Improved Chloroplast Count and Indirect Organogenesis of Indian Pea ( Pisum Sativum L.) Cv. Ageta 6

An effective callus-dependent indirect organogenesis protocol was standardized in recalcitrant Indian pea cv. Ageta 6 using cotyledonary node explant. This research highlighted the positive action of different polyamines (PA) such as spermidine (SPD), spermine (SPM), and putrescine (PUT) along with different plant growth regulators (PGRs) such as 2, 4-dichlorophenoxyacetic acid (2, 4-D), 6-benzyladenine (BA), and 1-naphthaleneacetic acid (NAA) on the enhancement of peas indirect organogenesis with callus induction, multiple shooting, and rooting parameters. The callus induction experiment demonstrated, 30 mg/L of PUT and 3 mg/L of 2, 4-D supplemented callus induction media has produced the highest green and white, friable pea callus induction with the highest explant response (46.26%). In the course of shoot multiplication study from the callus pieces of pea revealed that the highest multiple shooting efficiency (12.66 shoots/callus piece) under 20 mg/L of SPD and 1.5 mg/L BA treatments along with the highest explant response (53.66%) and shoot length (3.93 cm /shoots). The highest shoot elongation (6.20 cm length/shoots) was achieved under 1 mg/L GA 3 enriched shoot elongation media with maximum explant response (70%) even without polyamine assistance. The improved rooting analysis demonstrated, the highest root induction (10.86 roots/shoots) under 25 mg/L of PUT along with 0.6 mg/L of NAA supplemented rooting media with maximum root length (3.90 cm/shoots) and explant response (40%). Rooted plantlets were hardened effectively with a survival rate of 92%. The maximum callus induction (46.26%), shoot multiplication (12.66 shoot/callus piece), and root induction (10.86% root/shoot) were recorded in pea plant by the application of exogenous polyamines, which was approximately two to three-fold higher than the regular PGRs assisted indirect pea regeneration. The RAPD and SCoT molecular marker analysis justified the somoclonal variations free genomic steadiness in regenerated plants. Simultaneously, the polyamines-assisted regenerated pea plants showed three-fold improved photosynthetic (chlorophyll a, b and carotenoid) and two-fold improved antioxidant (DPPH, H 2 O 2 , and NO assay) profiles. Laser scanning confocal microscopy captured that the PA-assisted pea regenerants had a tremendous upsurge in the digit of precise chloroplasts compared to the control plant.