美国黄松组织培养不定根诱导的研究

以GD、SH和1／2SH基本培养基对美国黄松不定芽进行不定根的诱导。试验结果表明基本培养基的种类对不定芽形成不定根起主要作用。在1／2SH培养基上附加0．5mg／L的NAA不定根的诱导率为3．3％。试验首次在离体培养条件下,以美国黄松种胚为外植体获得了再生小植株。     

Microarray analyses of gene expression in Japanese flounder Paralichthys olivaceus leucocytes during monogenean parasite Neoheterobothrium hirame infection

In this study, the gene expression patterns of peripheral blood leucocytes (PBL) from Japanese flounder Paralichthys olivaceus were analyzed during the course of monogenean parasite Neoheterobothrium hirame infection in order to select candidates for molecular biomarkers of infection. cDNA microarray analysis was performed to compare the gene expression patterns of PBL between infected and non-infected fishes. Among the 797 genes analyzed, 45 genes (5.6%) changed their expression levels. These genes included specific and non-specific immune-related genes (matrix metalloproteinase[MMP]-9, MMP-13, leukotriene B4 receptor, CD20 receptor, MHC [major histocompatibility complex] Class I, MHC Class II beta-chain, immunoglobulin light chain and immunoglobulin heavy chain). Significant up- and down-regulation of some unknown genes was also observed. Several candidates for infection-marker genes were selected for further study. These genes included MMP-9, MMP-13, leukotriene b4 receptor, CD20 receptor, immunoglobulin heavy chain, immunoglobulin light chain and unknown genes coded as B613, E25, LB3(8), WE2(3), WE8-18R and WF12-18R.     

Mechanisms of primordium formation during adventitious root development from walnut cotyledon explants

 In walnut (Juglans regia L.), an otherwise difficult-to-root species, explants of cotyledons have been shown to generate complete roots in the absence of exogenous growth regulators. In the present study, this process of root formation was shown to follow a pattern of adventitious, rather than primary or lateral, ontogeny: (i) the arrangement of vascular bundles in the region of root formation was of the petiole type; (ii) a typical root primordium was formed at the side of the procambium within a meristematic ring of actively dividing cells located around each vascular bundle; (iii) the developing root apical meristem was connected in a lateral way with the vascular bundle of the petiole. This adventitious root formation occurred in three main stages of cell division, primordium formation and organization of apical meristem. These stages were characterized by expression of LATERAL ROOT PRIMORDIUM-1 and CHALCONE SYNTHASE genes, which were found to be sequentially expressed during the formation of the primordium. Activation of genes related to root cell differentiation started at the early stage of primordium formation prior to organization of the root apical meristem. The systematic development of adventitious root primordia at a precise site gave indications on the positional and biochemical cues that are necessary for adventitious root formation. Received: 30 July 1999 / Accepted: 16 February 2000     

Genome-wide expression analyses of gene regulation during early development of Dictyostelium discoideum

Using genome-wide microarrays, we recognized 172 genes that are highly expressed at one stage or another during multicellular development of Dictyostelium discoideum. When developed in shaken suspension, 125 of these genes were expressed if the cells were treated with cyclic AMP (cAMP) pulses at 6-min intervals between 2 and 6 h of development followed by high levels of exogenous cAMP. In the absence of cAMP treatment, only three genes, carA, gbaB, and pdsA, were consistently expressed. Surprisingly, 14 other genes were induced by cAMP treatment of mutant cells lacking the activatable adenylyl cyclase, ACA. However, these genes were not cAMP induced if both of the developmental adenylyl cyclases, ACA and ACR, were disrupted, showing that they depend on an internal source of cAMP. Constitutive activity of the cAMP-dependent protein kinase PKA was found to bypass the requirement of these genes for adenylyl cyclase and cAMP pulses, demonstrating the critical role of PKA in transducing the cAMP signal to early gene expression. In the absence of constitutive PKA activity, expression of later genes was strictly dependent on ACA in pulsed cells.     

Microarray analysis of gene expression during early adipocyte differentiation

The molecular mechanisms that regulate cellular differentiation during development and throughout life are complex. It is now recognized that precise patterns of differentially expressed genes ultimately direct a particular cell toward a given lineage and many of these are regulated during the earliest stages of differentiation. Using a microarray-based expression analysis, we have examined gene expression profiles during the first 24 h of 3T3-L1 adipocyte differentiation. RNA was isolated at times 0, 2, 8, 16, and 24 h following stimulation of differentiation and hybridized in duplicate to high density Affymetrix microarray gene chips containing a series of 13,179 cDNA/expressed sequence tag (EST) probe sets. Two hundred and eighty-five cDNA/ESTs were shown to have at least a fivefold change in expression levels during this time course and both hierarchical and self-organizing map clustering analysis was performed to categorize them by expression profiles. Several genes known to be regulated during this time period were confirmed and Western blot analysis of the proteins encoded by some of the identified genes revealed expression profiles similar to their mRNA counterparts. As expected, many of the genes identified have not been examined in such a critical time period during adipogenesis and may well represent novel adipogenic mediators.     

Microarray analyses of peripheral blood cells identifies unique gene expression signature in psoriatic arthritis

Psoriatic arthritis (PsA) is a chronic and erosive form of arthritis of unknown cause. We aimed to characterize the PsA phenotype using gene expression profiling and comparing it with healthy control subjects and patients rheumatoid arthritis (RA). Peripheral blood cells (PBCs) of 19 patients with active PsA and 19 age- and sex-matched control subjects were used in the analyses of PsA, with blood samples collected in PaxGene tubes. A significant alteration in the pattern of expression of 313 genes was noted in the PBCs of PsA patients on Affymetrix U133A arrays: 257 genes were expressed at reduced levels in PsA, and 56 genes were expressed at increased levels, compared with controls. Downregulated genes tended to cluster to certain chromosomal regions, including those containing the psoriasis susceptibility loci PSORS1 and PSORS2. Among the genes with the most significantly reduced expression were those involved in downregulation or suppression of innate and acquired immune responses, such as SIGIRR, STAT3, SHP1, IKBKB, IL-11RA, and TCF7, suggesting inappropriate control that favors proin-flammatory responses. Several members of the MAPK signaling pathway and tumor suppressor genes showed reduced expression. Three proinflammatory genes--S100A8, S100A12, and thioredoxin--showed increased expression. Logistic regression and recursive partitioning analysis determined that one gene, nucleoporin 62 kDa, could correctly classify all controls and 94.7% of the PsA patients. Using a dataset of 48 RA samples for comparison, the combination of two genes, MAP3K3 followed by CACNA1S, was enough to correctly classify all RA and PsA patients. Thus, PBC gene expression profiling identified a gene expression signature that differentiated PsA from RA, and PsA from controls. Several novel genes were differentially expressed in PsA and may prove to be diagnostic biomarkers or serve as new targets for the development of therapies.     

Microarray and real-time PCR analyses reveal mating type-dependent gene expression in a homothallic fungus

Sordaria macrospora is a homothallic ascomycete which is able to form fertile fruiting bodies without a mating partner. To analyze the molecular basis of homothallism and the role of mating products during fruiting body development, we have deleted the mating type gene Smta-1 encoding a high-mobility group domain (HMG) protein. The ΔSmta-1 deletion strain is morphologically wild type during vegetative growth, but it is unable to produce perithecia or ascospores. To identify genes expressed under control of Smta-1, we performed a cross-species microarray analysis using Neurospora crassa cDNA microarrays hybridized with S. macrospora targets. We identified 107 genes that are more than twofold up- or down-regulated in the mutant. Functional classification revealed that 81 genes have homologues with known or putative functions. Comparison of array data from ΔSmta-1 with those from three phenotypically similar mutants revealed that only a limited set of ten genes is deregulated in all mutants. Remarkably, the ppg2 gene encoding a putative lipopeptide pheromone is 500-fold down-regulated in the ΔSmta-1 mutant while in all other sterile mutants this gene is up-regulated. Electronic Supplementary Material Supplementary material is available for this article at and is accessible for authorized users.     

Inhibition of adventitious root development in apple rootstocks by cytokinin is based on its suppression of adventitious root primordia formation

Cytokinin (CK) inhibits adventitious root (AR) formation in stem cuttings. Little is known, however, about the mechanism underlying the inhibitory effect. In this study, 2 mg l^?1 of exogenous 6‐benzyl adenine (6‐BA) was administered to 3 and 7‐day‐old apple rootstocks ‘M.26’ cuttings (3 and 7 days 6‐BA) by transferring them from a rooting medium containing indole‐3‐butanoic acid to the medium containing 6‐BA. Anatomical and morphological observations revealed that the exogenous application of 6‐BA inhibited primordia formation in the 3 days 6‐BA but not the 7 days 6‐BA group. The concentration of auxin (IAA), the ratios of IAA/CK and IAA/abscisic acid were lower in 3 days 6‐BA than in 7 days 6‐BA. Expression analysis of genes known to be associated with AR formation was also analyzed. In the 3 days 6‐BA group, high level of CK inhibited the synthesis and transport of auxin, as a result, low endogenous auxin level suppressed the auxin signaling pathway genes, as were other AR development and cell cycle related genes; all of which had an inhibitory impact on AR primordium formation. On the contrary, low CK level in the 7 days 6‐BA, reduced the inhibitory impact on auxin levels, leading to an upregulated expression of genes known to promote AR primordia formation. Collectively, our data indicated that 3–7 days is the time period in which AR primordia formation occurs in cuttings of ‘M.26’ and that the inhibition of AR development by CK is due to the suppression of AR primordia development over 3–7 days period after culturing in rooting medium.     

Multilocus structure in Pinus contorta Dougl.

We studied isozyme variation at 21 loci in 66 populations from three subspecies of Pinus contorta Dougl.; 35 in spp. latifolia, 20 in spp contorta and 11 in spp. murrayana. The objectives were to assess gametic disequilibria and multilocus structure. There was considerable differentiation of allele frequencies at 19 polymorphic loci across the 66 populations and within the subspecies. Allele frequencies at many loci correlated with geographic variables. Genetic variability varied considerably among populations within subspecies but the subspecies means were similar. The mean number of polymorphic loci and the mean heterozygosity over 19 polymorphic loci were, respectively, 13 and 0.194 in latifolia, 12 and 0.196 in murrayana, and 12 and 0.180 in contorta. The mean heterozygosity correlated with longitude and altitude across the 66 populations and with latitude in latifolia. Gametic disequilibria were evident in 40 populations; 29 in latifolia, eight in murrayana and three in contorta. Gametic disequilibria correlated with latitude across the 66 populations and with longitude in latifolia. The single-locus F _ST averaged 0.0339 in latifolia, 0.0567 in murrayana, and 0.0764 in contorta. The multilocus F _STM was 0.1227 in latifolia, 0.2926 in murrayana, and 0.3328 in contorta. Multilocus Wahlund and founder effects, migration patterns, and natural selection, probably played significant roles in generating and maintaining the multilocus genetic structure in P. contorta in general and the subspecies latifolia in particular.     

CLAVATA1-LIKE, a leucine-rich-repeat protein receptor kinase gene differentially expressed during adventitious caulogenesis in Pinus pinaster and Pinus pinea

Molecular cloning and characterization of a CLAVATA1-LIKE gene in Pinus pinaster and Pinus pinea is reported. CLAVATA1 is a well-characterized gene in Arabidopsis integral to the balance between primordial differentiation and meristem proliferation. Currently, it is not known if the Arabidopsis model of in vitro caulogenesis is applicable to conifers. In this work, we study the putative role of the CLAVATA1-LIKE gene during caulogenic induction in cotyledons of P. pinea and P. pinaster after exposure to benzyladenine. Expression analysis showed that the gene was differentially expressed during the first phases of adventitious caulogenesis in cotyledons from both species, suggesting that CLAVATA1-LIKE may play a role in caulogenesis in conifers. A binary vector carrying CLAVATA1-LIKE promoter driven GFP:GUS expression was constructed. Results of genetic transformation showed GUS activity during somatic embryogenic mass proliferation and embryo maturation.     

Dihydroconiferyl alcohol in developing xylem of Pinus contorta

Dihydroconiferyl alcohol was identified by GC/MS as an endogenous constituent of developing xylem in Pinus contorta ssp. latifolia. Its role in regulating wood formation in P. contorta is discussed.