RESUMEN
Fern spores and pollen are haploid plant germplasm of microscopic nature that can be used to regenerate full plants through germination (fern spores) or to fertilize seed-bearing plants through breeding programs (pollen). Due to their short life span in conventional storage (i.e., dry at -20 °C), the use of cryopreservation has been indicated for long-term ex situ conservation. While fern spores of most species and pollen from many seeded plants tolerate desiccation and can be stored dry at liquid nitrogen temperatures, some pollen is desiccation sensitive, and cryopreservation protocols require controlled drying and cooling and some level of cryoprotection. In this chapter we describe the cryopreservation process for fern spores used in the Millennium Seed Bank of Royal Botanic Gardens, Kew, including some details of the fern spores harvest and cleaning methods. In addition, two protocols for pollen cryopreservation are described, one generic for desiccation-tolerant pollen that can be used for multiple species and one specific for a desiccation sensitive pollen (Zea mays).
Asunto(s)
Técnicas de Cultivo de Célula/métodos , Criopreservación/métodos , Crioprotectores/farmacología , Helechos/citología , Polen/citología , Esporas/citología , Venas Umbilicales/citología , Proliferación Celular , Células Cultivadas , Helechos/efectos de los fármacos , Polen/efectos de los fármacos , Esporas/efectos de los fármacos , Venas Umbilicales/efectos de los fármacosRESUMEN
Seeds are one of the preferable and most used sources of germplasm for the ex situ preservation of plant genetic resources. They are generally stored dry at -20 °C in seed banks following international standards. However, some seeds do not tolerate drying and/or storage at -20 °C, or present short lifespans at these conditions. For them cryopreservation is indicated for long-term preservation. When seeds tolerate desiccation (i.e., orthodox seeds), they can be dried at about 32 ± 3% relative humidity at 18 °C and stored in the vapor phase of liquid nitrogen. This is the method followed in the Millennium Seed Bank of the Royal Botanic Gardens, Kew, for wild species with short lifespans in the standard conditions of seed banks. When seeds do not tolerate desiccation (i.e., recalcitrant seeds) or their tolerance to desiccation and/or -20 °C storage is limited (i.e., intermediate seeds), drying and cooling procedures must be adjusted, and often, cryoprotection is also required. Some methods are detailed for diverse species of temperate and tropical origin.