Canine thyrotropin beta-subunit gene: cloning and expression in Escherichia coli, generation of monoclonal antibodies, and transient expression in the Chinese hamster ovary cells

The gene encoding the mature β subunit of canine thyroid stimulating hormone (cTSHβ) was cloned, sequenced and expressed in Escherichia coli and in Chinese hamster ovary (CHO) cells, and monoclonal antibodies against the recombinant cTSHβ purified from E. coli were generated. The gene fragment that encodes mature TSHβ was cloned from the canine genomic DNA by direct polymerase chain reaction (PCR) using primers that were designed based on the consensus sequences from other species. The resulting 891 basepairs (bp) of genomic DNA consisted of two coding exons of the canine TSHβ gene and an intron of 450 bp. The two exons, which encode the mature cTSHβ subunit, was joined together by an overlap PCR and was expressed in E. coli as 6×His-tagged protein. The purified recombinant cTSHβ with a molecular weight of about 15 kDa was recognized by the polyclonal antibodies prepared against the native canine TSH in Western blot. Monoclonal antibodies were raised against the purified cTSHβ and subsequently characterized. For transient expression in CHO cells that are permanently transfected with the bovine common gene, a 60-oligonucleotide signal peptide coding sequence was added to the 5′ end of the cTSHβ gene before it was cloned into the mammalian expression vector pRSV and used to transfect CHO cells. The medium from these transfected cells, presumably containing the bovine and canine TSHβ in heterodimeric confirmation, exhibited TSH bioactivity as indicated by the stimulation of cAMP production in the cultured FRTL-5 thyrocytes.     

Effect of different water supply on plant growth and fruit quality of Lycopersicon esculentum

It is well known from earlier work that water stress and salinity results in depressed plant growth and high fruit quality of tomato (e.g. increased sugar and acid levels), but generally is associated with a low marketable fruit yield. In the present work we investigated whether even a small reduction in water supply (without visible symptoms of water stress) also results in a high fruit quality together with high marketable fruit proportions. To characterize fruit quality sugars (glucose and fructose), titratable acids, odour-active aroma volatiles and vitamin C were investigated. Tomato plants (Lycopersicon esculentum Mill. cv Vanessa) were grown in soil and with the onset of fruit development water supply was varied (70% and 50% water capacity). In the treatment with lower water supply plant growth, and in particular the number of fruit settings were depressed and the sugar and vitamin C concentrations in the fruits were significantly increased, especially during fruit ripening. Furthermore, with lower water supply the concentrations of titratable acids and of C₆ aldehydes (hexanal, (Z)-3-hexenal and (E)-2-hexenal) were significantly increased in the red fruits. Fruit growth was identical in both treatments. The higher levels of sugars, titratable acids, aroma volatiles and vitamin C are responsible for the higher fruit quality under conditions of lower water supply. Since not all fruits of the well watered plants became mature, the marketable yield in both treatments was rather similar and hence, together with a higher fruit quality in the treatment with lower water supply, high proportions of marketable fruits can be harvested.     

The use of the ingrowth core method for measuring root production of arable crops — influence of soil conditions inside the ingrowth core on root growth

The ingrowth core method can be used to measure root gross growth (i.e. root production). A mesh bag filled with root free soil is buried into the root zone. After about 14 days, the bag is pulled out and root length inside the core can be determined. An objection against this method is the inability to obtain the same soil conditions inside the bag as outside, which can result in different root growth pattern in the ingrowth core compared to the bulk soil. To study this, mesh bags were buried in a stand of oilseed rape and were filled with soil at different nitrate, phosphate, moisture, and bulk density levels. Results showed that root growth was only influenced by a high nitrate content and a high soil density in the cores, which resulted in higher and lower root length densities (RLD), respectively. In a long‐term ingrowth experiment similar root length densities in the cores and in the bulk soil were measured, indicating that there were no root growth enhancing or impeding conditions inside the ingrowth cores. The conclusion is drawn, that the ingrowth core method gives reliable results, provided the N content and the soil density inside the bags are comparable to the bulk soil.     

Allometric above-belowground biomass equations for Nothofagus pumilio (Poepp. & Endl.) natural regeneration in the Chilean Patagonia

? In the present study, allometric biomass equations were developed for Nothofagus pumilio natural regeneration relating foliage, stem and branches (aboveground), roots (belowground), and total biomass to basal diameter and total height, based on destructive measurements of 390 naturally regenerated seedlings and saplings.

? Basal diameter was the most important independent variable in all equations and accounted for more than 88% of the variability of the different biomass components. The addition of height as a second independent variable slightly improved the predictions.

? The best-fit biomass components equations that were based on combinations of basal diameter and height as independent variables had adjusted R ² values between 0.80 and 0.95 and a mean percent standard error between 21.3% and 26.6%.

? Based on the best-fit biomass equations and the natural regeneration development in a 14-years chronosequence in forests managed under shelterwood cuts, the total biomass varied from 0.9 Mg ha^?1 (0.5 Mg ha^?1 above and 0.4 Mg ha^?1 belowground) for the primary forest, before the shelterwood cuts, to 19.5 Mg ha^?1 (13.6 Mg ha^?1 above and 5.9 Mg ha^?1 belowground) 14 years after the seed cut. In the same period, carbon storage varied, from 0.5 Mg ha^?1 to 9.8 Mg ha^?1.

     

Towards a classification and an understanding of developmental relationships of vertebral body malformations in Atlantic salmon (Salmo salar L.)

Vertebral column malformations in Atlantic salmon occur under farming conditions and in wild specimens and are commonly diagnosed by X-ray. The literature uses varied terms to describe malformations, and often, different terms refer to similar phenomena or similar terms describe unrelated pathologies. A coherent classification of spine defects is lacking. This shortcoming complicates the comparison of studies, makes it difficult to link particular malformations to particular causes and hampers the assessment of the prevalence of spinal deformities.Expanding available systems to categorise skeletal malformations in different teleost species, we propose 20 types of salmon vertebral column malformations that are repetitively observed under farming conditions. As vertebral column deformities are usually diagnosed by X-ray, the classification of deformity types is based on radiological observations that have been obtained over the past 10 years from more than 5000 specimens. The proposed deformity types should be identifiable independent from the type (analogue, digital) and the quality of radiographs. We propose nine types of compression and fusion-related deformities, four types of abnormal radiotranslucent or radiopaque vertebral bodies, three types of spine curvatures, three types of symmetry deviations and displacement of vertebral bodies and one type of severe multiple malformations. Based on the suggested categories we discuss the relationships between types of vertebral body malformations. Several types of malformations involve the development of heterotopic cartilage, a pathology that is possibly linked to mechanical overload and accelerated growth. Refined categories for vertebral body malformations should help link particular types of malformations to particular causes. It should also help find out which deformities represent final stages and which deformities represent transitory stages.     

Resistance of aquaculture net cage materials to biting by Atlantic Cod (Gadus morhua)

Cod bite on aquaculture net cages has resulted in damages like frayed netting and holes, which in part can explain why cultured cod have escaped more frequently than salmon over the last years. We describe damages found on various netting materials subjected to cod bite through field experiments at commercial cod farms. Further, a method to test local cod bite resistance of traditional netting structures is suggested and initial results from a test jig prototype are given. Results from field experiments indicated that cod may have been attracted by types of netting that made it possible to draw filaments out of the twine, while stiff, coated netting structures and thick filaments showed no sign of bite damage during the test period. We concluded that netting materials for cod aquaculture must be resistant to cod bite or be repellent or uninteresting for cod. Based on the present findings, the better choice among the traditional netting materials seemed to be hard-laid netting materials, preferably with a primer that glues the filaments together.     

Starch deposition in storage organs and the importance of nutrients and external factors

Starch is an important agricultural product deposited in vegetative and reproductive storage organs (sinks) of various crop species. Starch yield may in some cases be limited by photosynthesis, i.e. source-limited. This is particularly true for starch synthesized in potato tubers. Here, the physiological sink is characterized by a symplastic phloem unloading path. In reproductive storage tissues (seeds), however, photosynthates must pass the apoplast on their path from phloem unloading to the storage cell. In cereal grains, phloem unloading of sucrose and poslunloading processes rather than photosynthesis may thus control starch synthesis (sink-limited). Various limiting steps along the path of photosynthate movement from the phloem to the storage cells are considered. The primary organic carbon for starch synthesis is sucrose. Sucrose delivered to the storage cell is metabolized to UDPglucose and fructose by means of sucrose synthase activity. Concerning sucrose breakdown the role of cell-wall bound invertase is not well defined. Competition for UDPglucose consumed for growth or storage may be a crucial process in photosynthate partitioning. High starch yields of crops require an undisturbed growth of the sink organ and an optimal filling of sink amyloplasts with starch. The most important form of organic carbon imported into amyloplasts of storage organs (cereal grain and potato tuber) and used for starch synthesis is glucose 1-phosphate. It is still to be clarified whether the rate of glucose 1-phosphate absorption has a direct impact on starch yield. In cereals, the total amount of starch accumulated depends significantly on the duration of grain filling. Ample nutrient and water supply at the post-anthesis stage prolongs the period of grain filling and hence favours starch production. High temperature reduces the activity of soluble starch synthase with negative consequences for starch accumulation. The biochemical and physiological implications of these stress factors are discussed. Recently, successful transgenic manipulations of starch synthesis in crop plants have increased starch yield.