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<p>Cryopreservation of in vitro-derived explants offers a secure and cost-effective strategy for long-term germplasm conservation, particularly for species unsuitable for conventional seed storage. <i>Momordica dioica</i> Roxb. ex Willd., a nutritionally and medicinally important dioecious cucurbit, faces conservation constraints due to poor seed viability under low-temperature storage and lack of true-to-type regeneration from seed-derived plants. This study established an efficient in vitro propagation protocol and, for the first time, a sex-specific shoot-tip cryopreservation method for <i>M. dioica</i>. Aseptic cultures of both female (SKNF) and male (RMDSG-2) genotypes were initiated from nodal explants of sprouted tubers. Optimal bud induction occurred on Murashige and Skoog (MS) medium with 0.88 µM 6-benzylaminopurine (BAP). Maximum proliferation was achieved in the female genotype (7.8 shoots per explant) on MS with 0.88 µM BAP and 200&#xa0;mg L⁻<sup>1</sup> casein hydrolysate, and in the male (7.4 shoots per explant) on MS with 2.22 µM BAP. Rooting was best on half-strength MS medium supplemented with 4.92 µM indole-3-butyric acid (IBA). For long-term storage, droplet-vitrification was standardized using in vitro-derived axillary shoots. The highest post-thaw survival and regeneration occurred in explants pre-cultured with 0.3&#xa0;M sucrose for 24&#xa0;h, followed by 20&#xa0;min exposure each to loading solution and chilled PVS2, with the maximum response recorded in female genotype (73.40% and 65.95%) compared to the male genotype (57.86%, 51.76%). Genetic fidelity of regenerated plants was assessed using eight simple sequence repeat (SSR) and expressed sequence tag (EST)- SSR primers. All primers produced monomorphic banding patterns with no detectable polymorphism at the analysed loci, confirming genetic uniformity with the mother plants. The developed system enables efficient clonal propagation and reliable long-term, genotype-specific conservation of <i>M. dioica</i>, aiding preservation of its valuable genetic diversity.</p>

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Droplet vitrification-based cryopreservation for long-term conservation of spiny gourd (Momordica dioica Roxb. ex Willd.)

  • 期刊论文

  • 发表日期：

    2026年3月30日

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  • Vartika Srivastava,
  • K. Pradheep,
  • Jitendra Kumar Tiwari,
  • N. N. Prajapati,
  • Padmavati Ganpat Gore,
  • Anuradha Agrawal,
  • S. K. Kaushik,
  • Monika Singh,
  • Rakesh Singh,
  • Anju Mahendru Singh,
  • Gyanendra Pratap Singh

摘要

Cryopreservation of in vitro-derived explants offers a secure and cost-effective strategy for long-term germplasm conservation, particularly for species unsuitable for conventional seed storage. Momordica dioica Roxb. ex Willd., a nutritionally and medicinally important dioecious cucurbit, faces conservation constraints due to poor seed viability under low-temperature storage and lack of true-to-type regeneration from seed-derived plants. This study established an efficient in vitro propagation protocol and, for the first time, a sex-specific shoot-tip cryopreservation method for M. dioica. Aseptic cultures of both female (SKNF) and male (RMDSG-2) genotypes were initiated from nodal explants of sprouted tubers. Optimal bud induction occurred on Murashige and Skoog (MS) medium with 0.88 µM 6-benzylaminopurine (BAP). Maximum proliferation was achieved in the female genotype (7.8 shoots per explant) on MS with 0.88 µM BAP and 200 mg L⁻1 casein hydrolysate, and in the male (7.4 shoots per explant) on MS with 2.22 µM BAP. Rooting was best on half-strength MS medium supplemented with 4.92 µM indole-3-butyric acid (IBA). For long-term storage, droplet-vitrification was standardized using in vitro-derived axillary shoots. The highest post-thaw survival and regeneration occurred in explants pre-cultured with 0.3 M sucrose for 24 h, followed by 20 min exposure each to loading solution and chilled PVS2, with the maximum response recorded in female genotype (73.40% and 65.95%) compared to the male genotype (57.86%, 51.76%). Genetic fidelity of regenerated plants was assessed using eight simple sequence repeat (SSR) and expressed sequence tag (EST)- SSR primers. All primers produced monomorphic banding patterns with no detectable polymorphism at the analysed loci, confirming genetic uniformity with the mother plants. The developed system enables efficient clonal propagation and reliable long-term, genotype-specific conservation of M. dioica, aiding preservation of its valuable genetic diversity.