Seed survival in Chilean Nothofagus in response to desiccation and storage

Nothofagus alpina, N. obliqua, N. glauca, N. leonii, N. dombeyi and N. pumilio seeds exhibited consistent, albeit slight, sensitivity to extreme desiccation, but nevertheless maintained viability at low moisture contents and cool temperatures (-10 degrees to -20 degrees C) over 2 years. Nothofagus alpina, N. obliqua, N. glauca, N. leonii and N. dombeyi conformed to the seed viability equation of Ellis and Roberts; sensitivity of longevity to temperature was quantitatively similar to that of crop seeds, sensitivity to moisture was somewhat less, and a low-moisture-content limit to the equation was detected at 4.8% moisture content in hermetic storage at 65 degrees C, and possibly similar moisture contents at 30-40 degrees C. These five species show orthodox seed storage behaviour. Therefore, ex-situ conservation of these Nothofagus species in seed banks is possible, but the quality of seed lots collected requires attention. Seed storage behaviour was not defined in N. pumilio: initial seed quality was poor and loss of viability was detected over 2 years at 0 degrees, -10 degrees and -20 degrees C at 2.7% moisture content, but not at 5.2%. The results confirm that the economy of nature in seed storage physiology extends to forest tree seeds, but the repeated observation of reduced sensitivity of longevity to moisture in forest tree seeds requires further investigation.

Role of polyhydroxybutyrate and glycogen as carbon storage compounds in pea and bean bacteroids

Rhizobium leguminosarum synthesizes polyhydroxybutyrate and glycogen as its main carbon storage compounds. To examine the role of these compounds in bacteroid development and in symbiotic efficiency, single and double mutants of R. legumosarum bv. viciae were made which lack polyhydroxybutyrate synthase (phaC), glycogen synthase (glgA), or both. For comparison, a single phaC mutant also was isolated in a bean-nodulating strain of R. leguminosarum bv. phaseoli. In one large glasshouse trial, the growth of pea plants inoculated with the R. leguminosarum bv. viciae phaC mutant were significantly reduced compared with wild-type-inoculated plants. However, in subsequent glasshouse and growth-room studies, the growth of pea plants inoculated with the mutant were similar to wildtype-inoculated plants. Bean plants were unaffected by the loss of polyhydroxybutyrate biosynthesis in bacteroids. Pea plants nodulated by a glycogen synthase mutants or the glgA/phaC double mutant, grew as well as the wild type in growth-room experiments. Light and electron micrographs revealed that pea nodules infected with the glgA mutant accumulated large amounts of starch in the II/III interzone. This suggests that glycogen may be the dominant carbon storage compound in pea bacteroids. Polyhydroxybutyrate was present in bacteria in the infection thread of pea plants but was broken down during bacteroid formation. In nodules infected with a phaC mutant of R. leguminosarum bv. viciae, there was a drop in the amount of starch in the II/III interzone, where bacteroids form. Therefore, we propose a carbon burst hypothesis for bacteroid formation, where polyhydroxybutyrate accumulated by bacteria is degraded to fuel bacteroid differentiation.

Seed development, maturation and storage behaviour of Mimusops elengi L

Mass maturity (end of the seed-filling phase) occurred at about 72 days after flowering (DAF) in developing seeds of Mimusops elengi, at which time seed moisture content had declined to about 55%. The onset of ability to germinate was detected at 56 DAF and seeds showed 98% germination by 84 DAF. Tolerance of desiccation to 10% moisture content was first detected at 70 DAF and was maximal by 84 DAF. Delaying collection by a further 14 days to 98 DAF, when fruits began to be shed, reduced seed viability, particularly for seeds first dried to 10% moisture content. Hence the best time for seed collection appears to be about 14 days before fruits shed. In a separate investigation with six different seed lots, desiccation below about 8-12% moisture content reduced viability (considerably in some lots). The viability of dry seeds (below about 10% moisture content) stored hermetically was reduced at cool temperatures (5 degrees C and below), and none survived storage at sub-zero temperatures. The results suggest that Mimusops elengi shows intermediate seed storage behaviour and that the optimal hermetic seed storage environment is about 10% moisture content at 10 degrees C, while short-term, moist, aerated storage at high (40%) moisture content is also feasible.

Successful long-term ultra dry storage of seed of 15 species of Brassicaceae in a genebank: variation in ability to germinate over 40 years and dormancy

Seed of 15 species of Brassicaceae were stored hermetically in a genebank (at -5 degrees C to -10 degrees C with c. 3% moisture content) for 40 years. Samples were withdrawn at intervals for germination tests. Many accessions showed an increase in ability to germinate over this period. due to loss in dormancy. Nevertheless, some dormancy remained after 40 years' storage and was broken by pre-applied gibberellic acid. The poorest seed survival occurred in Hormatophylla spinosa. Even in this accession the ability to germinate declined by only 7% between 1966 and 2006. Comparison of seeds from 1966 stored for 40 years with those collected anew in 2006 from the original sampling sites, where possible, showed few differences, other than a tendency (7 of 9 accessions) for the latter to show greater dormancy. These results for hermetic storage at sub-zero temperatures and low moisture contents confirm that long-term seed storage can provide a successful technology for ex situ plant biodiversity conservation.

Suitability of cryopreservation for the long-term storage of rare and endangered plant species: a case history for Cosmos atrosanguineus

The suitability of cryopreservation for the secure, long-term storage of the rare and endangered species Cosmos atrosanguineus was investigated. Using encapsulation/dehydration of shoot tips in alginate strips, survival rates of up to 100 % and shoot regeneration of up to 35 % were achieved. Light and electron microscopy studies indicated that cellular damage to some regions of the shoot tip during the freeze/thaw procedure was high, although cell survival in and around the meristematic region allowed shoot tip regeneration. The genetic fingerprinting technique, amplified fragment length polymorphisms (AFLPs), showed that no detectable genetic variation was present between material of C. atrosanguineus at the time of initiation into tissue culture and that which had been cryopreserved, stored in liquid nitrogen for 12 months and regenerated. Wearied plantlets that were grown under glasshouse conditions exhibited no morphological variation from non-frozen controls. (C) 2003 Annals of Botany Company.

Juvenile-mature wood transition in pine: correlation between wood properties and candidate gene expression profiles

There is a strong desire to exploit transcriptomics data from model species for the genetic improvement of non-model crops. Here, we use gene expression profiles from the commercial model Pinus taeda to identify candidate genes implicated in juvenile-mature wood transition in the non-model relative, P. sylvestris. Re-analysis of 'public domain' SAGE data from xylem tissues of P. taeda revealed 283 mature-abundant and 396 juvenile-abundant tags (P < 0.01), of which 70 and 137, respectively matched to genes with known function. Based on sequence similarity, we then isolated 16 putative homologues of genes that in P. taeda exhibited widest divergence in expression between juvenile and mature samples. Candidate expression levels in P. sylvestris were almost invariably differential between juvenile and mature woody tissue samples among two cohorts of five trees collected from the same seed source and selected for genetic uniformity by genetic distance analysis. However, the direction of differential expression was not always consistent with that described in the original P. taeda SAGE data. Correlation was observed between gene expression and juvenile-mature wood anatomical characteristics by OPLS analysis. Four candidates (alpha-tubulin, porin MIP1, lipid transfer protein and aquaporin like protein) apparently had greatest influence on the wood traits measured. Speculative function of these genes in relation to juvenile-mature wood transition is briefly explored. Thus, we demonstrate the feasibility of exploiting SAGE data from a model species to identify consistently differentially expressed candidates in a related non-model species.

Role of polyhydroxybutyrate and glycogen as carbon storage compounds in pea and bean bacteroids

Rhizobium leguminosarum synthesizes polyhydroxybutyrate and glycogen as its main carbon storage compounds. To examine the role of these compounds in bacteroid development and in symbiotic efficiency, single and double mutants of R. legumosarum bv. viciae were made which lack polyhydroxybutyrate synthase (phaC), glycogen synthase (glgA), or both. For comparison, a single phaC mutant also was isolated in a bean-nodulating strain of R. leguminosarum bv. phaseoli. In one large glasshouse trial, the growth of pea plants inoculated with the R. leguminosarum bv. viciae phaC mutant were significantly reduced compared with wild-type-inoculated plants. However, in subsequent glasshouse and growth-room studies, the growth of pea plants inoculated with the mutant were similar to wildtype-inoculated plants. Bean plants were unaffected by the loss of polyhydroxybutyrate biosynthesis in bacteroids. Pea plants nodulated by a glycogen synthase mutants or the glgA/phaC double mutant, grew as well as the wild type in growth-room experiments. Light and electron micrographs revealed that pea nodules infected with the glgA mutant accumulated large amounts of starch in the II/III interzone. This suggests that glycogen may be the dominant carbon storage compound in pea bacteroids. Polyhydroxybutyrate was present in bacteria in the infection thread of pea plants but was broken down during bacteroid formation. In nodules infected with a phaC mutant of R. leguminosarum bv. viciae, there was a drop in the amount of starch in the II/III interzone, where bacteroids form. Therefore, we propose a carbon burst hypothesis for bacteroid formation, where polyhydroxybutyrate accumulated by bacteria is degraded to fuel bacteroid differentiation.

Cellulose microfibril angle in the cell wall of wood fibres

The term microfibril angle (MFA) in wood science refers to the angle between the direction of the helical windings of cellulose microfibrils in the secondary cell wall of fibres and tracheids and the long axis of cell. Technologically, it is usually applied to the orientation of cellulose microfibrils in the S2 layer that makes up the greatest proportion of the wall thickness, since it is this which most affects the physical properties of wood. This review describes the organisation of the cellulose component of the secondary wall of fibres and tracheids and the various methods that have been used for the measurement of MFA. It considers the variation of MFA within the tree and the biological reason for the large differences found between juvenile (or core) wood and mature (or outer) wood. The ability of the tree to vary MFA in response to environmental stress, particularly in reaction wood, is also described. Differences in MFA have a profound effect on the properties of wood, in particular its stiffness. The large MFA in juvenile wood confers low stiffness and gives the sapling the flexibility it needs to survive high winds without breaking. It also means, however, that timber containing a high proportion of juvenile wood is unsuitable for use as high-grade structural timber. This fact has taken on increasing importance in view of the trend in forestry towards short rotation cropping of fast grown species. These trees at harvest may contain 50% or more of timber with low stiffness and therefore, low economic value. Although they are presently grown mainly for pulp, pressure for increased timber production means that ways will be sought to improve the quality of their timber by reducing juvenile wood MFA. The mechanism by which the orientation of microfibril deposition is controlled is still a matter of debate. However, the application of molecular techniques is likely to enable modification of this process. The extent to which these techniques should be used to improve timber quality by reducing MFA in juvenile wood is, however, uncertain, since care must be taken to avoid compromising the safety of the tree.

Cryopreservation of cocoa (Theobroma cacao L.) somatic embryos for long-term germplasm storage

Cryopreservation using encapsulation-dehydration was developed for the long-term conservation of cocoa (Theobroma cacao L.) germplasm. Survival of individually encapsulated somatic embryos after desiccation and cryopreservation was achieved through optimization of cryoprotectants (abscisic acid (ABA) and sugar), duration of osmotic and evaporative dehydration, and embryo development stage. Up to 63% of the genotype SPA4 early-cotyledonary somatic embryos survived cryopreservation following 7 days preculture with 1 M sucrose and 4 h silica exposure (16% moisture content in bead). This optimized protocol was successfully applied to three other genotypes, e.g. EET272, IMC14 and AMAZ12, with recovery frequencies of 25, 40 and 72%, respectively (but the latter two genotypes using 0.75 M sucrose). Recovered SPA4 somatic embryos converted to plants at a rate of 33% and the regenerated plants were phenotypically comparable to non-cryopreserved somatic embryo-derived plants.

Stability of plasmid profile of highly virulent races of Xanthomonas campestris pv. malvacearum during storage and subculturing

Four races of Xanthomonas campestris pv. mal-vacearum (Xcm) viz. races 23, 27 and 32 (isolated from Gossypium hirsutum) and race 23b (from Gossypium barbadense) were studied. The plasmid profile of the natural isolates showed four plasmids in races 23 and 23b (ca. 60, 40, 23, 8.2 kb), five in race 27 (ca. 60, 40, 23, 8.2 and 3.7 kb) and six in race 32 (ca. 60, 40, 23, 8.2, 3.7 and 1.6 kb). Continuously sub-cultured laboratory isolates of the Xcm races resulted in the loss of all but two plasmids, ca. 60 and 40 kb in size. When the laboratory isolates were passed through cotton (Gossypium hirsutum), they regained certain plasmids so that four plasmids were found in race 23 and 23b (ca. 60, 40, 23 and 8.2 kb), five in race 27 (ca. 60, 40, 23, 8.2 and 3.7 kb) and six in race 32 (ca. 60, 40, 23, 8.2, 3.7 and 1.6 kb), which was more or less similar to the original isolates. The isolates recovered from cotton maintained their plasmid profile (except for minor changes in the miniplasmids) after storage for six months at -70degreesC in 50% glycerol. It is suggested that plasmid profiles among highly virulent races of Xcm are unstable during repeated sub-culturing at room temperature, resulting in rapid loss of some plasmids. However, when the cultures were sub-cultured and stored at -70degreesC the plasmid profile was fairly stable except for the miniplasmids (ca. 3.7 and 1.6 kb).