The purpose of our research was to establish a protocol for the in vitro culture of Viburnum treleasei, a rare and endangered taxon with high ornamental potential endemic to the Azores islands. The surface sterilization of the explants was better achieved with a pretreatment of 0.1% (w/v) Benomyl for 2 h followed by 0.2% (w/v) HgCl2 for 10 min with agitation. Shoot tips were the most efficient explants for shoot development and single-node segments for proliferation. Woody plant medium (WPM) was adequate for all micropropagation stages. For culture establishment and shoot development, a hormone-free medium was adequate, whereas a 1.1 μM N6-benzyl adenine medium supplement was more efficient for shoot multiplication. Elongation and rooting could be carried out on a 1.3 μM 1-naphthaleneacetic acid-supplemented medium. Acclimatization of in vitro-produced plantlets was achieved after 1 month with a success rate of 50%. This in vitro propagation procedure will be useful for the conservation of Viburnum treleasei through production of morphologically true-to-type plants, allowing the recovery of depleted natural populations. Chemical names used: N6-benzyl adenine (BA); 1-naphthaleneacetic acid (NAA); HgCl2 (mercury bichloride).
Mónica Moura, Maria Irene Candeias and Luís Silva
Orlanda Cristina Barros Moreira, José Martins, Luís Silva and Mónica Moura
Prunus azorica is an Azorean endemic tree considered as a priority species for conservation. It is important as a laurel forest component, particularly at medium altitude, and as a food source for the endangered bird Pyrrhula murina. The best conditions for seed germination were investigated after removal of the outer layers of the fruit by determining the effect of 1) using stones or seeds; 2) stratification regime (six treatments and a control); 3) incubation temperature (four alternating temperature regimes); and 4) gibberellic acid concentration (three levels). This resulted in a fully factorial design with 168 (2 × 7 × 4 × 3) treatments with three replicates per treatment and 25 seeds per replicate. Cumulative germination percentages were determined at the end of the trial. Globally, there was a significant effect of endocarp removal (49% germination with seeds and 15% with stones). Both for stones and seeds, there was a significant effect of incubation temperature, stratification regime, and growth regulator concentration. Stones attained a maximum germination of ≈80% under several stratification treatments including cold (4 °C) or warm (20 °C) followed by cold and at 10/5 °C without the addition of a growth regulator. Seeds attained a maximum germination of greater than 90% without stratification at 10/5 or 15/10 °C without the addition of a growth regulator. During the stratification process, germination occurred only for seeds, particularly for longer treatments, for example, those corresponding to 3 or more months of stratification, including warm followed by cold (75% to 80%) or cold alone (77%). According to seed morphology and germination results, the seed appears to have a non-deep physiological dormancy. Seeds of P. azorica can thus be efficiently germinated after endocarp removal at temperatures of 10/5 or 15/10 °C with a daily light period of 12 hours. This protocol allowed producing hundreds of viable seedlings that were used in the reforestation of a laurel forest stand in a LIFE project.
Salvador Guzmán-González, Pedro Valadez-Ramírez, Rosa-Edith Robles-Berber, Laura Silva-Rosales and José-Luis Cabrera-Ponce
Biolistic genetic transformation of plants with viral genes is a method for controlling plant virus diseases; however, optimization of the particle bombardment parameters according to the transformation system is a key factor for an appropiate transgene expression and, therefore, a stronger resistance mechanism in transgenic plants. In order to optimize biolistic parameters, somatic papaya (Carica papaya L.) cv. Maradol embryo masses were bombarded with the CAMBIA 1301 plasmid construction that contains the coat protein gene (CP) of the papaya ringspot virus isolate of Colima, Mexico, driven by the double constitutively CaMV 35S promoter and flanked for the GUS and hygromycin (hpt) resistance genes. Particle bombardment protocol was carried out using the Helios™ Gene Gun device (BioRad) and the manufacturer's instruction manual. Helium pressure (50, 100, and 150 psi) and gold particle size (0.6, 1.0, and 1.6 μm) were evaluated. Five days after bombardment, somatic embryo clusters were used for GUS transient expression and, during 2 months, were selected into 50, 75, and 150 mg·L-1 hygromycin-containing media to its later CP-PCR detection. Results showed that 50 psi and 1.0 μm were the two optimal values for the assayed analyses. This is the first report of genetic transformation of papaya using the Helios™ Gene Gun device as a new tool compared to conventional PDS-1000/He.
Valdinar Ferreira Melo, Edvan Alves Chagas, Raphael Henrique da Silva Siqueira, Olisson Mesquita de Souza, Luís Felipe Paes de Almeida, Diogo Francisco Rossoni, Pollyana Cardoso Chagas and Carlos Abanto-Rodríguez
Camu-camu is an native species in domestication processes. Therefore, studies related to the root system are necessary to evaluate the best cultivation practices for an orchard. Two trials were conducted, one with nitrogen (N) and the other with potassium (K), at doses of 0, 40, 80, 160, and 320 kg·ha–1. Root distribution was determined using nondestructive analyses in which two-dimensional (2D) root images were obtained from trenches under the plants’ canopy. Variables included width (measured in cubic millimeters), area (measured in square millimeters), and length (measured in millimeters, and were analyzed using Safira software (version 2010, Embrapa, Brazil). To have better spatial visualization of variable distribution in the soil profile, data were analyzed using the ordinary kriging technique with the geoR software package and R software (version 2016). Both N and K doses influenced positively the camu-camu root system with regard to length, volume, and area. Better root distribution was verified with 80 kg·ha−1 N and 160 kg·ha−1 K doses. The nondestructive analysis via 2D images allowed sound characterization of root spatial distribution.