Anther culture for haploid production in guava (Psidium guajava L.)

Development of haploid and Doubled Haploids through in vitro androgenesis can greatly accelerate the process of achieving homozygosity in plants. In tree species like citrus, papaya, neem, mulberry, custard apple and apple, doubled haploids have been produced successfully. In current study we tested androgenesis in three phenotypically variable genotypes of guava viz. Allahabad Safeda, Purple Guava and Punjab Pink. Anthers with microspores at mid to late uninucleate stage corresponding to unopened flower bud stage IV on N6 basal media containing 1% PVP showed maximum callus induction of 16.3±0.27% in Allahabad Safeda followed by 10±0.41% in Punjab Pink and 4.6±0.27% in Purple Guava. The stereomicroscopic evaluation of callus generating anthers indicated calli being produced from microspores. The callus induction frequency was enhanced to 27.7±1.18% by 3–7 days pre treatment of cold (4 oC) plus 8% mannitol to unopened flower bud stage IV. 77% calli on basal N6 nourished with liquid media (N6 + 0.5 mg/L BAP + 0.5 mg/L Kin + 1% PVP + 3% sucrose + 50 ml/L Coconut water) in the mother test tube without mechanical disturbance for 1 month with few calli exhibited secondary structures. Ovaries excised from unopened flower bud stage IV also underwent callusing but after 4 months of in vitro culturing on 1% PVP containing basal MS media underwent necrosis. Callus originating from ovaries underwent necrosis after one week of sub culturing on MS media fortified with BAP (0.2mg/L) and Kinetin (0.2mg/L). Success of doubled haploid in guava can result into generating homozygous inbreds and such inbreds upon crossing shall show high hybrid vigour. Genome sequencing of homozygous guava plant can help generate the complete and unambiguous genome assembly as in apple, citrus and grapes.