Characterization of inflorescence-predominant chitinase gene in Metroxylon sagu via differential display

Chitinase is an enzyme that catalyzes the degradation of chitin, commonly induced upon the attack of pathogens and other stresses. A cDNA (MsChi1) was isolated from Metroxylon sagu and expressed predominantly in the inflorescence tissue of M. sagu, suggesting its role in developmental processes. The chitinase cDNA was detected and isolated via differential display and rapid amplification of cDNA ends (RACE). Primers specific to M. saguchitinase were used as probes to amplify the 3′-end and 5′-end regions of chitinase cDNA. Transcript analysis showed that chitinase is expressed in inflorescence and meristem tissues but was not detected in the leaf tissue. Sequence analysis of amplified cDNA fragments of 3′-end and 5′-end regions indicated that the chitinase cDNA was successfully amplified. The M. saguchitinase cDNA isolated was approximately 1,143 bp long and corresponds to 312 predicted amino acids. Alignments of nucleotide and amino acid have grouped this chitinase to family 19 class I chitinase

Regulation of somatic embryo development in Norway spruce (Picea abies)

Somatic embryos can be used for vegetative propagation of genetically superior material. Additionally, the system offers unique opportunities to study embryology. Somatic embryogenesis has been induced in many coniferous species and the process is similar between species. A common problem is, however, that not all embryogenic cell lines contain embryos that can mature. We are using Norway spruce (Picea abies) as a model system to study how the development of the conifer somatic embryo is regulated. Different cell lines contain embryos that have reached a specific developmental stage, which is reflected in the pattern of extracellular proteins and arabinogalactan proteins present. The embryo morphology is partially regulated by proteins. Proteins identified as having a putative regulatory effect on embryo development include chitinases, peroxidases, zeamatin-like proteins and arabinogalactan proteins.Régulation du développement des embryons somatiques chez l'épicéa. Les embryons somatiques peuvent être utilisés pour la propagation végétative de matériels améliorés génétiquement. Cette méthode offre en plus des opportunités uniques pour étudier l'embryologie. L'embryogenèse somatique a été induite chez de nombreuses espèces de conifères selon un processus identique parmi ces espèces. Cependant il faut noter que, de façon générale, toutes les lignées cellulaires embryogènes ne donnent pas forcément des embryons capables de maturation. Nous utilisons l'épicéa (Picea abies) en tant que système modèle pour étudier comment est régulé le développement des embryons somatiques de conifères. Les différentes lignées cellulaires contiennent des embryons qui ont atteint un stade spécifique de leur développement, ce dernier étant caractérisé par un profil de protéines extracellulaires et la présence de protéines arabinogalactiques. La morphologie des embryons est en partie régulée par des protéines ; parmi celles qui ont été identifiées comme ayant un pouvoir régulateur, il y a des chitinases, des péroxydases, des protéines de type zéamatine et des protéines arabinogalactiques

Effect of cryopreservation on growth of different elite embryogenic cell lines of norway spruce (Picea Abies (L.) Karst.)

Callus from seven different elite embryogenic cell lines of Norway spruce originated from the controlled crossing conducted by Skogforsk (Forest Research Institute of Sweden) have been cryopreserved in liquid nitrogen using the slow freezing method with sorbitol (0.2 and 0.4 M) and DMSO (5% v/v) as cryoprotectants. Cooloing of samples was performed in a programmable freezer Cryo Med 7452. The cryovials were inserted at +4°C and frozen at -0.3°C/min to -16°C. After 15 min at -16°C, to avoid rapid cooling and crystal formation in the cells, the cryovials were further cooled to -35°C at -0.3°C/min. At the end of the freezing program the cryovials were transferred and stored in a CBS V1500 liquid nitrogen storage unit at -196°C. After thawing and plating on proliferation medium, the recovered cells showed a short lag phase, after which they continued to grow. Lags in growth were also observed after the transfer to pre-maturation and maturation media. Somatic embryo maturation and plantlet regeneration occurred in all the tested embryogenic lines