Influence of the cryopreservation technique, recovery medium and genotype on genetic stability of mint cryopreserved shoot tips

摘要

This study compared the effect of the cryopreservation protocol, the genotype and the plant growth regulators used in the recovery media on the genetic stability of mint shoot tips. Shoot tips of two micropropagated genotypes of mint were cryopreserved either using an encapsulation-dehydration or droplet-vitrification protocol. Three recovery media were tested, all of them MS based although with different plant growth regulators: (1) 0.5 mg L-1 6-benzylaminopurine (BAP); (2) 0.5 mg L-1 6-dimethylallylamino-purine (2iP) + 0.1 mg L-1 alpha-naphthalene acetic acid (NAA); and (3) 0.5 mg L-1 BAP + 0.1 mg L-1 NAA. DNA was extracted from three different types of samples: leaves from shoots, callus at the base of shoots and callus. RAPD markers were used to assess the genetic stability. One of the genotypes, 'MEN 198', showed higher percentage of stable samples than the other one, 'MEN 186' (97 vs. 42 %; considering all media, protocol and type of explant). The material recovered after droplet-vitrification showed higher stability compared to the encapsulation-dehydration recovered samples: 99 versus 87 % in 'MEN 198', and 80 versus 24 % in 'MEN 186'. There were not large differences in the genetic stability obtained from the three types of samples in 'MEN 196' (85-100 %), while in 'MEN 186' calli were more unstable (53, 44 and 30 % stability for leaves, basal callus and callus, respectively). The effect of the recovery medium on the stability of the samples was noticeable in the more unstable types of explants but not in leaves.