Quantitative genetics of growth and development in Populus. I. A three-generation comparison of tree architecture during the first 2 years of growth

One approach to gain an insight into the genetics of tree architecture is to make use of morphologically divergent parents and study their segregating progeny in the F₂ and backcross (B₁) generations. This approach was chosen in the present study in which material of a three-generation pedigree growing side by side in a replicated plantation, was analyzed. The pedigree included Populus trichocarpa (T) and P. deltoides (D) parents, their F₁ and F₂ hybrids and their B₁ hybrids to the D parent. The trees were grown in the environment of the T parent and measured for the first 2 years of growth. Nine quantitative traits were studied at the stem, branch and leaf levels of tree architecture, in which the original parents differed. Strong F₁ hybrid vigor relative to the better parent (T) was expressed in growth and its components. Most quantitative traits in the F₂ and B₁ hybrids were intermediate between the T and D parents but displayed a wide range of variation due to segregation. The results from the analysis of variance indicated that all morphometric traits were significantly different among F₂ and B₁ clones, but the B₁ hybrids were more sensitive to replicates than the F₂. Broad-sense heritabilities (H ²) based on clonal means ranged from moderately high to high (0.50–0.90) for the traits studied, with H ² values varying over age. The H ² estimates reflected greater environmental noise in the B₁ than in the F₂, presumably due to the greater proportion of maladaptive D alleles in those hybrids. In both families, sylleptic branch number and length, and leaf size on the terminal, showed strong genetic correlations with stem growth. The large divergence between the two original parents in the traits studied, combined with the high chromosome number in Populus (2n=38), makes this pedigree well suited for the estimation of the number of quantitative trait loci (QTLs) underlying quantitative variation by Wright's biometric method (1968). Variation in several traits was found to be under the control of surprisingly few major QTLs: 3–4 in 2nd-year height and diameter growth, a single QTL in stem diameter/height ratio.     

A general suppressor of RNA polymerase I,II and III mutations in Saccharomyces cerevisiae

The pem locus, which is responsible for the stable maintenance of the low copy number plasmid R100, contains the pemK gene, whose product has been shown to be a growth inhibitor. Here, we attempted to isolate mutants which became tolerant to transient induction of the PemK protein. We obtained 20 mutants (here called pkt for PemK tolerance), of which 9 were temperature sensitive for growth. We analyzed the nine mutants genetically and found that they could be classified into three complementation groups, pktA, pktB and pktC, which corresponded to three genes, ileS, gltX and asnS, encoding isoleucyl-, glutamyl- and asparaginyl-tRNA synthetases, respectively. Since these aminoacyl-tRNA synthetase mutants did not produce the PemK protein upon induction at the restrictive temperature, these mutants could be isolated because they behaved as if they were tolerant to the PemK protein. The procedure is therefore useful for isolating temperature-sensitive mutants of aminoacyl-tRNA synthetases.     

The genetic resolution of juvenile canopy structure and function in a three-generation pedigree of Populus

 A genetic approach to the understanding of tree architecture is to cross trees of contrasting features and to study their segregating F₂ progenies. For this purpose, members of a 3-generation pedigree, combining Populus trichocarpa, P. deltoides, and their F₁ and F₂ offspring, were grown side by side in a clonally replicated plantation. At 2 and 3 years of growth, tree architecture was analyzed at the stem, branch, and leaf levels. In all generations, proleptic branches were more numerous, longer, and had more and larger leaves than sylleptics initiated in the same year. The analysis of variance revealed significant genotypic effects on growth, branch and leaf biometrics in the F₂ family, with broad-sense heritabilities (H²) ranging from 0.50 to 0.80 for most traits. For branch and leaf traits, the H² values were found to vary among branch types and crown positions. In year 2, the degree of genetic control was stronger for sylleptics than proleptics and for upper than lower crown positions. These patterns were followed in year 3, except that H² values were more a function of position within crown, as a consequence of increased competition among trees. The genetic correlations between branch/leaf morphology and stem growth were also a function of branch type and crown position. Generally, traits on proleptics or at upper positions were more tightly correlated with height growth, whereas those on sylleptics or at lower positions, with basal area growth. By year 3, proleptic traits showed increased genetic correlations with both height and radial growth. The implications of these results for the construction of ideotypes are discussed. Received: 1 December 1995     

Physical map of the Methanobacterium thermoautotrophicum Marburg chromosome.

A physical map of the Methanobacterium thermoautotrophicum Marburg chromosome was constructed by using pulsed-field gel electrophoresis of restriction fragments generated by NotI, PmeI, and NheI. The order of the fragments was deduced from Southern blot hybridization of NotI fragment probes to various restriction digests and from partial digests. The derived map is circular, and the genome size was estimated to be 1,623 kb. Several cloned genes were hybridized to restriction fragments to locate their positions on the map. Genes coding for proteins involved in the methanogenic pathway were located on the same segment of the circular chromosome. In addition, the genomes of a variety of thermophilic Methanobacterium strains were treated with restriction enzymes and analyzed by pulsed-field gel electrophoresis. The sums of the fragment sizes varied from 1,600 to 1,728 kb among the strains, and widely different macrorestriction patterns were observed.     

An essential and specific subunit of RNA polymerase III (C) is encoded by gene RPC34 in Saccharomyces cerevisiae.

The RPC34 gene of Saccharomyces cerevisiae was cloned by immunological screening, using antibodies raised against the C34 polypeptide of the RNA polymerase III (C). This single copy gene was located near the centromere of chromosome XIV. It included a coding sequence of 317 amino acids that strictly matched two internal oligopeptides of C34. This polypeptide is a specific component of RNA polymerase III, with no significant homology to any other RNA polymerase subunit known so far. It is an essential subunit, since inactivation by deletion or nonsense mutations led to a recessive lethal phenotype. Moreover, a partially blocked mutant, rpc34-F297, had a reduced tRNA synthesis in vivo but no detectable effect on 5 S RNA synthesis. The latter phenotype was observed for all conditionally defective RNA polymerase III mutants isolated so far.     

A high-throughput media design approach for high performance mammalian fed-batch cultures

An innovative high-throughput medium development method based on media blending was successfully used to improve the performance of a Chinese hamster ovary fed-batch medium in shaking 96-deepwell plates. Starting from a proprietary chemically-defined medium, 16 formulations testing 43 of 47 components at 3 different levels were designed. Media blending was performed following a custom-made mixture design of experiments considering binary blends, resulting in 376 different blends that were tested during both cell expansion and fed-batch production phases in one single experiment. Three approaches were chosen to provide the best output of the large amount of data obtained. A simple ranking of conditions was first used as a quick approach to select new formulations with promising features. Then, prediction of the best mixes was done to maximize both growth and titer using the Design Expert software. Finally, a multivariate analysis enabled identification of individual potential critical components for further optimization. Applying this high-throughput method on a fed-batch, rather than on a simple batch, process opens new perspectives for medium and feed development that enables identification of an optimized process in a short time frame.     

The Effect of Aerobic Exercise on Intrahepatocellular and Intramyocellular Lipids in Healthy Subjects

Background

Intrahepatocellular (IHCL) and intramyocellular (IMCL) lipids are ectopic lipid stores. Aerobic exercise results in IMCL utilization in subjects over a broad range of exercise capacity. IMCL and IHCL have been related to impaired insulin action at the skeletal muscle and hepatic level, respectively. The acute effect of aerobic exercise on IHCL is unknown. Possible regulatory factors include exercise capacity, insulin sensitivity and fat availability subcutaneous and visceral fat mass).

Aim

To concomitantly investigate the effect of aerobic exercise on IHCL and IMCL in healthy subjects, using Magnetic Resonance spectroscopy.

Methods

Normal weight, healthy subjects were included. Visit 1 consisted of a determination of VO_2max on a treadmill. Visit 2 comprised the assessment of hepatic and peripheral insulin sensitivity by a two-step hyperinsulinaemic euglycaemic clamp. At Visit 3, subcutaneous and visceral fat mass were assessed by whole body MRI, IHCL and IMCL before and after a 2-hours aerobic exercise (50% of VO_2max) using ¹H-MR-spectroscopy.

Results

Eighteen volunteers (12M, 6F) were enrolled in the study (age, 37.6±3.2 years, mean±SEM; VO_2max, 53.4±2.9 mL/kg/min). Two hours aerobic exercise resulted in a significant decrease in IMCL (−22.6±3.3, % from baseline) and increase in IHCL (+34.9±7.6, % from baseline). There was no significant correlation between the exercise-induced changes in IMCL and IHCL and exercise capacity, subcutaneous and visceral fat mass and hepatic or peripheral insulin sensitivity.

Conclusions

IMCL and IHCL are flexible ectopic lipid stores that are acutely influenced by physical exercise, albeit in different directions.

Trial Registration

ClinicalTrial.gov {"type":"clinical-trial","attrs":{"text":"NCT00491582","term_id":"NCT00491582"}}NCT00491582     

Culture of Echinococcus multilocularis metacestodes: an alternative to animal use

This article reviews the use of an in vitro culture model for the maintenance and proliferation of Echinococcus multilocularis metacestodes and the formation of protoscoleces. This model has been used to identify and characterize parasite molecules involved in host-parasite interactions, and is a suitable tool to perform in vitro drug-screening assays. The development of a simple and easy-to-handle assay to determine the effects of drugs on parasite viability, without the need for time-consuming animal experimentation, has opened the way for larger-scale in vitro drug screening.     

Bacterial diversity and community composition in the chemocline of the meromictic alpine Lake Cadagno as revealed by 16S rDNA analysis

Using different techniques of molecular biology we investigated the bacterial diversity of the chemocline of the meromictic Lake Cadagno. Cloning of a total community 16S rDNA PCR product and subsequent screening with a combination of amplified ribosomal DNA restriction analysis and temporal temperature gradient gel electrophoresis (TTGE) analysis revealed that 30 of 47 randomly selected clones were unique. Partial sequencing and comparative analysis indicated a high bacterial diversity dominated by the gamma-Proteobacteria (33.3%). Most of these rDNA clone sequences were not closely related to any 16S rDNA sequence in the database. In a second approach, the TTGE pattern from an environmental sample was compared with the migration of the cloned 16S rDNA fragments. Four clone types were identified on the environmental pattern by excising and sequencing comigrating bands, three of which were well represented in the library: two Chromatiaceae species and one sequence affiliated with the Desulfobulbus assemblage. Using the fluorescent in situ hybridization technique we essentially confirmed the results of the cloning experiments and the TTGE analysis.     

Genetic variation and productivity of Populus trichocarpa and its hybrids. X. Trait correlations in young black cottonwood from four river valleys in Washington

 A common-garden study of Populus trichocarpa Torr. & Gray was established in spring 1986 with 128 clones collected from sites along two mesic (Hoh and Nisqually) and two xeric (Dungeness and Yakima) river valleys in Washington. Two replicate plantations, one in Puyallup and the other in Wenatchee, Wash., were established with this material. Over 2 years data were taken on stem growth, leaf/crown characters, spring/autumn phenology, and the incidence of Melampsora occidentalis leaf rust. Combining clones from all four sources, correlation/regression analyses were used to examine clonal stability of traits between test sites and trait relationships with stem growth; broad-sense heritabilities (H ²) and genetic correlations revealed the genetic strength of these traits. At Puyallup, many leaf/crown traits predicted stem growth moderately to very well (r ²>0.50), e.g., total leaf area (TLA) to diameter gave an r ² of 0.91 and current-terminal leaf size, of 0.79. Some regressions were quadratic, suggesting a threshold level in a trait (e.g., leaf size) beyond which stem growth levels off. Upper-crown TLA was more closely related to height than TLA of the lower sylleptics, but the reverse was true for diameter. A decline in r ² values from upper to lower crown positions was sharper for correlations of TLAs with height than with diameter. Thus, leaf area allocation seems to differentially affect stem growth. When autumn leaf fall (LF) and rust incidence (R) were regressed with growth, r ² values ranged from 0.58 to 0.71, but those of spring flush (SF) were only 0.10 to 0.12. Early LF and high R, both negatively affecting growth, had a strong geographic component as it occurred mainly on lower-elevation Yakima clones. At Wenatchee, field conditions were harsher and microsites more variable, so trait/growth relationships were weaker. Genetic correlations with growth revealed similar trends as phenotypic analyses. Unlike leaf/crown traits, clonal scores of LF, SF, and R were fairly stable across the two test sites (r ²: 0.58–0.80). These traits also showed strong genetic control (H ²: 0.96–0.98). The trait/growth relationships as well as trait stability within clones have implications for selecting clonal stock in poplar culture and conservation. Received: 23 February 1998 / Accepted: 7 May 1998