RESUMO
In this study, we successfully cryopreserved cotyledonary somatic embryos of diploid and triploid Arachis pintoi cytotypes using the encapsulation-dehydration technique. The highest survival rates were obtained when somatic embryos were encapsulated in calcium alginate beads and precultured in agitated (80 rpm) liquid establishment medium (EM) with daily increasing sucrose concentration (0.50, 0.75, and 1.0 M). The encapsulated somatic embryos were then dehydrated with silica gel for 5 h to 20% moisture content (fresh weight basis) and cooled either rapidly (direct immersion in liquid nitrogen, LN) or slowly (1 degree C per min from 25 degree C to -30 degree C followed by immersion in LN). Beads were kept in LN for a minimum of 1 h and then were rapidly rewarmed in a 30 degree C water-bath for 2 min. Finally, encapsulated somatic embryos were post-cultured in agitated (80 rpm) liquid EM with daily decreasing sucrose concentration (0.75 and 0.5 M) and transferred to solidified EM. Using this protocol, we obtained 26% and 30% plant regeneration from cryopreserved somatic embryos of diploid and triploid cytotypes. No morphological abnormalities were observed in any of the plants regenerated from cryopreserved embryos and their genetic stability was confirmed with 10 isozyme systems and nine RAPD profiles.
Assuntos
Arachis/embriologia , Criopreservação/métodos , Alginatos/química , Dessecação , Ácido Glucurônico/química , Ácidos Hexurônicos/química , Dióxido de Silício/químicaRESUMO
A cDNA clone (peanut Gly-1) encoding for glycinin protein was isolated from the immature seeds (from yellow-1 maturity pods) and characterized. The clone spanning a total of 1836 bp, predicted protein of 529 amino acid residues with a calculated mass of 60,447.61 Da. Peanut Gly-1 sequence comparison shows high level of sequence homology with other two peanut glycinin (arachin) genes [Ara h3 (95 percent) and Ara h4 (94 percent)] and glycinin (legumin) genes of other legumes such as soybean, broad bean, French bean and pea etc., both at nucleotide (67 to 69 percent) and amino acid (60 to 63 percent) levels. The N- and C-terminals of peanut Gly-1 are highly conserved with other glycinin genes; central region of the gene possess three variable regions, which also show conservation with other glycinin genes. Peanut glycinin-1 gene deciphers 11S type A seed storage protein. Mapping for conserved domains indicate that Peanut Gly-1 consists of bi-cupin domain. RNA gel blot studies demonstrate that the gene expressed during embryo development that is transcriptionally activated early in embryogenesis (white pod maturity) and is repressed late in seed maturation (orange pod maturity stage). Peanut Gly-1 does not express in other tissues like leaf, stem, root, flower, pegs or post germinating seedlings.
Assuntos
Arachis/genética , Globulinas/genética , Arachis/embriologia , Sequência de Bases , Northern Blotting , Clonagem Molecular , Fabaceae/genética , Reação em Cadeia da Polimerase , Proteínas de Plantas/genética , Proteínas de SojaRESUMO
The effects of two methods of cryopreservation involving chemical vitrification and air desiccation) were studied on isolated embryonic axes of A. hypogaea. Vitrification with PVS2 and desiccation in a laminar flow cabinet resulted in high levels (70-90%) of whole plant recovery after cryopreservation. A desiccation protocol based on 1h exposure of explants to the air flow was successfully applied to six wild species of section Extranervosae, resulting in recovery levels of 70-90% after liquid nitrogen treatment.