In-vitro Propagation, Phytochemical-antioxidant Screening and Dna Fingerprinting of Papaya (Carica Papaya L.)

Abstract

Papaya (Carica papaya L.) is well known for its exceptional and medicinal properties throughout the world but one of the major problems is low genetic diversity of commercial genotypes which makes this crop plants more susceptible to pests and diseases. Therefore, in order to understand the genetic diversity, 20 SSR markers were used among a set of 25 papaya genotypes and a total of 51 alleles were detected with an average of 2.55 alleles/locus. The PIC values for all the polymorphic primers ranged from 0.073 to 0.943 with an average value of 0.637. Two primers, CP02 and CP13 showed highest PIC values (0.943 and 0.933, respectively). These 2 polymorphic primers can effectively be used in further molecular breeding programs and QTL mapping studies of papaya since they exhibited very high polymorphism over other loci. On the basis of dendogram, 2 varieties (Pusa Nanha and LV11) from different clusters were selected for phytochemical-antioxidant screening. In present study, the phyto-constituents of the two varieties were similar as they contained the same secondary metabolites in all the extracts. Results of the study also indicated that polar solvents (methanol, ethanol and aqueous) contained more phytochemicals than the non-polar once (n-hexane). The highest yield percent of Pusa Nanha and LV11 was found to be in aqueous extract (16.3% and 14.24%, respectively) followed by methanol extract (10.46% and 8.82%, respectively), ethanol extract (6.46% and 5.12%, respectively) and n-hexane extract (2.16% and 1.96%, respectively). The highest TPC value of Pusa Nanha and LV11 was found to be in methanolic extract (58.6±0.18 and 31.12±0.59 mg GAE/g, respectively), TFC was found to be maximum in n-hexane extract (31.5±0.2 and 26±1.08 mg QE/g, respectively) and higher free-radical scavenging activity was found in methanolic extract with an IC50 value of 4.464 mg/mL and 34.51 mg/L, respectively. Pusa Nanha was found to contain more antioxidant activity as compare to local variety 11. Therefore, an efficient protocol for callus induction (via leaf and petiole explants) and direct organogenesis (via lateral buds) was established. Minimum time span for callus induction and higher percent of explants produced callus was found to be in media containing 3.0 mg/L NAA + 0.5 mg/L Kin + 1.5 μM TDZ for both leaf (90%) and petiole (11%). Sterilant combination, 0.1% Bavistin (10 min) + 0.1% HgCl2 (2 min) + 4% NaOCl (5 min) + 70% EtOH (1 min) was found to be the most effective way for higher explants survival with minimum contamination. Shaking the lateral buds in 300 mg/L rifampicin before inoculation posses a great role to obtain higher survival %. Maximum number of shoots and shoot length was observed in media having 30 g/L sucrose while culturing lateral buds. Higher percent of surviving explants (66.66±2.02 %), highest number of shoots per explant (11.33±0.33) and longest shoot (2.30±0.13 cm) was recorded in media containing 1.0 mg/L BAP + 0.2 mg/L NAA. The maximum percent of explants produced shoot (37.33±1.45 %) was observed in media containing 0.5 mg/L BAP + 0.2 mg/L NAA. Half strength MS + 1.5 mg/L IBA was found to be the best for rooting parameters such as minimum time (15.66±0.88 days), maximum percent of shoot produced root (49.33±1.45), maximum number of roots (4.00±0.57) and highest root length (1.50±0.05 cm). The plantlets with well-developed roots were successfully transplanted to plastic pots containing mixture of compost and soil with 30% survival rate after 15 days of transfer.