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- Lemm, Julie A, et al.
(författare)
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Template-dependent initiation of Sindbis virus RNA replication in vitro
- 1998
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Ingår i: Journal of Virology. - 0022-538X .- 1098-5514. ; 72:8, s. 6546-6553
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Tidskriftsartikel (refereegranskat)abstract
- Recent insights into the early events in Sindbis virus RNA replication suggest a requirement for either the P123 or P23 polyprotein, as well as mature nsP4, the RNA-dependent RNA polymerase, for initiation of minus-strand RNA synthesis. Based on this observation, we have succeeded in reconstituting an in vitro system for template-dependent initiation of SIN RNA replication. Extracts were isolated from cells infected with vaccinia virus recombinants expressing various SIN proteins and assayed by the addition of exogenous template RNAs. Extracts from cells expressing P123C>S, a protease-defective P123 polyprotein, and nsP4 synthesized a genome-length minus-sense RNA product. Replicase activity was dependent upon addition of exogenous RNA and was specific for alphavirus plus-strand RNA templates. RNA synthesis was also obtained by coexpression of nsP1, P23C>S, and nsP4. However, extracts from cells expressing nsP4 and P123, a cleavage-competent P123 polyprotein, had much less replicase activity. In addition, a P123 polyprotein containing a mutation in the nsP2 protease which increased the efficiency of processing exhibited very little, if any, replicase activity. These results provide further evidence that processing of the polyprotein inactivates the minus-strand initiation complex. Finally, RNA synthesis was detected when soluble nsP4 was added to a membrane fraction containing P123C>S, thus providing a functional assay for purification of the nsP4 RNA polymerase.
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- Olsson, P.A., et al.
(författare)
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Growth of arbuscular mycorrhizal mycelium in calcareous dune sand and its interaction with other soil microorganisms as estimated by measurement of specific fatty acids
- 1998
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Ingår i: Plant and Soil. - 0032-079X. ; 201:1, s. 9-16
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Tidskriftsartikel (refereegranskat)abstract
- Fatty acid analysis was used for determining the extent of the development of the external mycelium of AM fungi (mixed inoculum from a sand dune) growing from roots of Festuca rubra and Plantago lanceolata in calcareous dune sand. The plants were raised in chambers specially designed to permit the growth of AM mycelium in root-free compartments. In two separate experiments, growth of external mycelium in the root-free compartments was detected and the amount of mycelium was estimated using the indicator of AM fungal biomass, phospholipid fatty acid (PLFA) 16:1 omega 5. The results showed that the PLFA 16:1 omega 5 was suitable for estimating the mycelium emerging from the mixed inoculum obtained from the field roots of F: rubra and P lanceolata. The PLFA 16:1 omega 5 showed external mycelium to become established in the root-free compartments within a period of 3 weeks and the amount of mycelium to continue to increase at 6 and 9 weeks. Increases in neutral lipid fatty acid (NLFA) 16:1 omega 5 (indicator of storage lipids) over time were inconsistent between the two experiments, but appeared to follow patterns of sporulation in each experiment. In both experiments, the root-free compartment was colonised by saprophytic fungi to a greater extent in the case of non-mycorrhizal than of AM treatment, as indicated by an increase in PLFA 18:2 omega 6,9 (indicator of saprophytic fungi). The absence of an increase in the case of AM treatment indicates that AM fungal mycelium can negatively affect the growth of saprophytic fungi in this soil type. This result was, however, only weakly supported by measurements of ergosterol content. The analysis of bacteria specific PLFAs showed that bacterial biomass was not affected by the AM mycelium.
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