Advances in the cryopreservation of Coffea canephora seeds

Abstract

Currently, coffee germplasm is conserved in collections of live plants in the field (ex situ), which requires a lot of space and maintenance, there is a chance of an environmental disaster occurring. Therefore, it is necessary to study other conservation options. Cryopreservation is already an option for orthodox seeds, but for intermediate and recalcitrant seeds further studies are needed and the protocols used vary according to the species. So far, Coffea canephora Pierre seeds, when cryopreserved, have low survival and the biggest obstacles to the technique are the cooling and warming stages. Therefore, further studies are needed in order to increase survival after cryopreservation. This study aimed to evaluate the best time and temperature for warmingC. canephora Pierre seeds after being submitted to the cryopreservation stages, through physiological, biochemical, ultrastructural and molecular studies.For this, tests were carried out to better understand what happens in the seeds during the steps of the technique. The research was carried out at the Central Laboratory of Seed Analysis, of the Department of Agriculture of ESAL, UFLA, with seeds of C. canephora Pierre, cultivar Apoatã. The harvested fruits were mechanically hulledand the seeds were dried in silica gel or in a stationary dryer until a humidity of 20% (wb). After drying, the seeds were placed in mesh bags and immersed directly in liquid nitrogen.The seeds were removed and warmed in a water bath at different temperatures (40, 45, 50, 55, 60, 70ºC) for 1, 2 and 3 minutes. Germination, tetrazolium and electrical conductivity tests were carried out, in addition to biochemical, molecular and scanning electron microscopy evaluations. Warming at a temperature of 70ºC is harmful and should not be recommended for cryopreservation protocols for Coffea canephora Pierre seeds, regardless of the exposure time. Drying in silica gel, aiming at cryopreservation, increases the chances of success of the technique, when compared to drying in the dryer, for Coffea canephora Pierre seeds.Drying on silica gel to a water content of 20% (wb), followed by direct immersion in liquid nitrogen and warming at a temperature of 50ºC, for 2 minutes, in a water bath, ensured greater survival of the seeds.