Characterization of glucoannylaserom Asperigillus terreus 4

A strain of Aspergillus terreus 4 was found to show extracellular amylolytic activity and the amylase was identified as glucoamylase enzyme. The optimum temperature for the enzyme activity was 60% and it was stable at this temperature for 1 h. The enzyme was optimally active at pH 5.0 and stable between pH 3.0-8.0. Km values of glucoamylase for soluble starch, amylose and amylopectin were 5.9 mg/ml, 4.8 mg/ml and 2.6 mg/ml respectively.     

Genetic mapping,synteny, and physical location of two loci for Fusarium oxysporum f. sp. tracheiphilum race 4 resistance in cowpea [Vigna unguiculata (L.) Walp]

Fusarium wilt is a vascular disease caused by the fungus Fusarium oxysporum f.sp. tracheiphilum (Fot) in cowpea [Vigna unguiculata (L.) Walp]. In this study, we mapped loci conferring resistance to Fot race 4 in three cowpea RIL populations: IT93K-503-1 × CB46, CB27 × 24-125B-1, and CB27 × IT82E-18/Big Buff. Two independent loci which confer resistance to Fot race 4 were identified, Fot4-1 and Fot4-2. Fot4-1 was identified in the IT93K-503-1 (resistant) × CB46 (susceptible) population and was positioned on the cowpea consensus genetic map, spanning 21.57–29.40 cM on linkage group 5. The Fot4-2 locus was validated by identifying it in both the CB27 (resistant) × 24-125B-1 (susceptible) and CB27 (resistant) × IT82E-18/Big Buff (susceptible) populations. Fot4-2 was positioned on the cowpea consensus genetic map on linkage group 3; the minimum distance spanned 71.52–71.75 cM whereas the maximum distance spanned 64.44–80.23 cM. These genomic locations of Fot4-1 and Fot4-2 on the cowpea consensus genetic map, relative to Fot3-1 which was previously identified as the locus conferring resistance to Fot race 3, established that all three loci were independent. The Fot4-1 and Fot4-2 syntenic loci were examined in Glycine max, where several disease-resistance candidate genes were identified for both loci. In addition, Fot4-1 and Fot4-2 were coarsely positioned on the cowpea physical map. Fot4-1 and Fot4-2 will contribute to molecular marker development for future use in marker-assisted selection, thereby expediting introgression of Fot race 4 resistance into future cowpea cultivars.     

Mapping QTL for drought stress-induced premature senescence and maturity in cowpea [Vigna unguiculata (L.) Walp.]

Cowpea is an important crop for subsistence farmers in arid regions of Africa, Asia, and South America. Efforts to develop cultivars with improved productivity under drought conditions are constrained by lack of molecular markers associated with drought tolerance. Here, we report the mapping of 12 quantitative trait loci (QTL) associated with seedling drought tolerance and maturity in a cowpea recombinant inbred (RIL) population. One hundred and twenty-seven F₈ RILs developed from a cross between IT93K503-1 and CB46 were screened with 62 EcoR1 and Mse1 primer combinations to generate 306 amplified fragment length polymorphisms for use in genetic linkage mapping. The same population was phenotyped for maintenance of stem greenness (stg) and recovery dry weight (rdw) after drought stress in six greenhouse experiments. In field experiments conducted over 3 years, visual ratings and dry weights were used to phenotype drought stress-induced premature senescence in the RIL population. Kruskall–Wallis and multiple-QTL model mapping analysis were used to identify QTL associated with drought response phenotypes. Observed QTL were highly reproducible between stg and rdw under greenhouse conditions. Field studies confirmed all ten drought-response QTL observed under greenhouse conditions. Regions harboring drought-related QTL were observed on linkage groups 1, 2, 3, 5, 6, 7, 9, and 10 accounting for between 4.7 and 24.2% of the phenotypic variance (R ²). Further, two QTL for maturity (R ² = 14.4–28.9% and R ² = 11.7–25.2%) mapped on linkage groups 7 and 8 separately from drought-related QTL. These results provide a platform for identification of genetic determinants of seedling drought tolerance in cowpea. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

The International Barley Sequencing Consortium—At the Threshold of Efficient Access to the Barley Genome

Arabidopsis (Arabidopsis thaliana) tryptophan-proline-proline (WPP)-domain proteins, WPP1 and WPP2, are plant-unique, nuclear envelope-associated proteins of unknown function. They have sequence similarity to the nuclear envelope-targeting domain of plant RanGAP1, the GTPase activating protein of the small GTPase Ran. WPP domain-interacting tail-anchored protein 1 (WIT1) and WIT2 are two Arabidopsis proteins containing a coiled-coil domain and a C-terminal predicted transmembrane domain. They are required for RanGAP1 association with the nuclear envelope in root tips. Here, we show that WIT1 also binds WPP1 and WPP2 in planta, we identify the chaperone heat shock cognate protein 70-1 (HSC70-1) as in vivo interaction partner of WPP1 and WPP2, and we show that HSC70-1 interacts in planta with WIT1. WIT1 and green fluorescent protein (GFP)-WIT1 are targeted to the nuclear envelope in Arabidopsis. In contrast, GFP-WIT1 forms large cytoplasmic aggregates when overexpressed transiently in Nicotiana benthamiana leaf epidermis cells. Coexpression of HSC70-1 significantly reduces GFP-WIT1 aggregation and permits association of most GFP-WIT1 with the nuclear envelope. Significantly, WPP1 and WPP2 show the same activity. A WPP1 mutant with reduced affinity for GFP-WIT1 fails to decrease its aggregation. While the WPP-domain proteins act on a region of WIT1 containing the coiled-coil domain, HSC70-1 additionally acts on the C-terminal transmembrane domain. Taken together, our data suggest that both HSC70-1 and the WPP-domain proteins play a role in facilitating WIT1 nuclear envelope targeting, which is, to our knowledge, the first described in planta activity for the WPP-domain proteins.The cytoplasmic Ran GTPase activating protein RanGAP is critical to establishing a functional RanGTP/RanGDP gradient across the nuclear envelope (NE) and is associated with the outer surface of the NE in metazoan and higher plant cells (Matunis et al., 1996; Rose and Meier, 2001). Plant RanGAP1 association with the NE requires a plant-specific targeting domain, named the Trp-Pro-Pro (WPP) domain (Rose and Meier, 2001). Arabidopsis (Arabidopsis thaliana) WPP1 and WPP2 are small (155- and 180-amino-acid residues, respectively) plant-unique proteins of unknown function, which are similar to the WPP domain of RanGAP proteins. WPP1 and WPP2 are located in the cytoplasm, with a concentration at the NE (Patel et al., 2004). They are characterized by a 104-amino-acid-long WPP domain, predicted to consist of a β-strand and three α-helices and shown to be sufficient for NE targeting (Patel et al., 2004). They are also associated with cytoplasmic speckles most likely representing Golgi (Patel et al., 2005). Reduced expression of the WPP protein family causes decreased mitotic activity in roots of Arabidopsis, resulting in shortening of primary roots and decreased number of lateral roots (Patel et al., 2004). RanGAP1 association with the NE in the Arabidopsis root tip requires two families of NE-localized, plant-specific, WPP domain-interacting proteins (WPP domain-interacting protein [WIP] and WPP domain-interacting tail-anchored protein [WIT] families) that are characterized by the presence of a coiled-coil domain and a C-terminal predicted transmembrane domain (TMD; Xu et al., 2007; Zhao et al., 2008). Based on sequence analysis, both the WIP and WIT protein family were classified as putative tail-anchored (TA) proteins, proteins that associate with membranes posttranslationally (Borgese et al., 2003).The heat shock protein 70 family (HSP70) contains both heat-inducible and constitutively expressed members, called heat shock cognate proteins (HSC70). HSC70 chaperones assist in folding newly synthesized proteins (Bukau and Horwich, 1998), are involved in posttranslational translocation of secretory proteins across endoplasmic reticulum (ER) and mitochondrial membranes (Chirico et al., 1988; Deshaies et al., 1988), prevent irreversible aggregation of their substrates (Ngosuwan et al., 2003), and facilitate degradation of misfolded proteins (Meacham et al., 2001). Recently, mammalian HSC70 has also been implied in assisting the membrane insertion of a subset of TA proteins (Abell et al., 2007).The Arabidopsis genome encodes five different cytosolic HSP70s, three of which are expressed constitutively (HSC70-1, HSC70-2, and HSC70-3). While expressed in all organs, Hsc70-1 and Hsc70-2 expression levels are highest in leaves and Hsc70-3 in leaves and roots. All three genes can be further induced by heat shock and cold stress (Sung et al., 2001). Constitutive overexpression of Arabidopsis Hsc70-1 in transgenic plants leads to changes in growth and development, increases thermotolerance (Sung and Guy, 2003), and decreases the plant''s ability to respond to pathogen attack (Noel et al., 2007). Recently, specific interactions of HSC70-1 with SGT1 (for Suppressor of G2 allele of skp1; Noel et al., 2007) and HSC70-3 with turnip mosaic virus RNA-dependent RNA polymerase (Dufresne et al., 2008) were identified, suggesting a role of HSC70 in viral replication and pathogenesis. Both HSC70-1 and HSC70-3 can be detected in the nuclei and the cytoplasm of Nicotiana benthamiana epidermal cells (Noel et al., 2007; Dufresne et al., 2008).Here, we identified Arabidopsis HSC70-1 as an in vivo interaction partner of WPP1 and WPP2 and demonstrated that HSC70-1 associates with WIT1. Using transient expression in N. benthamiana, we show that when expressed at a high level, WIT1 accumulates in large fluorescent bodies in the cytoplasm that may represent aggregates. Upon coexpression in the same system, WPP1, WPP2, and HSC70-1 are all able to prevent the aggregation of overexpressed WIT1 and enable WIT1 association with the NE. While WPP-domain proteins act on a region of WIT1 containing the coiled-coil domain, HSC70-1 additionally acts on the C-terminal TMD. We propose that WPP1 and WPP2 play a chaperone-like role reflected in preventing the aggregation of the coiled-coil region of WIT1 and possibly other coiled-coil TA-type proteins, either in conjunction or independently of HSC70-type chaperones.     

Ecophysiology of Species with Distinct Leaf Morphologies: Effects of Plastic and Shadecloth Tree Guards

Ecological restoration using seedling tubestock is challenging under a Mediterranean-type climate of hot, dry summers. We investigated the ecophysiological effects of plastic tree guards and shadecloth tree guards during seedling establishment of four co-occurring tree species that differ in leaf morphology. Average temperature was 6.7°C higher in plastic guards than controls over a summer, with a maximum of 53.5°C compared to 47.9°C in controls. Light levels were 2-fold lower in both tree guard treatments relative to control. In spring, photosynthesis and specific leaf area were significantly elevated in shadecloth tree guards relative to other treatments. In summer, photosynthetic rate was significantly lower, and midday photochemical efficiency was significantly higher, in both tree guard treatments relative to controls. The effect of elevated temperature in plastic tree guards may partially explain our results of higher mortality of seedling in plastic tree guards. The relatively elevated spring photosynthesis of seedlings in shadecloth tree guards may partially explain the result of reduced mortality and increased growth in this treatment. We conclude that shadecloth tree guards create a microclimate more favorable for seedling establishment in a Mediterranean-type environment than plastic tree guards and control treatments. Our results may have wide applicability to the range of restoration settings where seedling tubestock is planted, except in environments where low temperature is limiting to plant growth.     

Understorey thinning and burning trials are needed in conservation reserves: The case of Tuart (Eucalyptus gomphocephala D.C.)

Summary Management interventions are needed to reverse the decline of Tuart (Eucalyptus gomphocephala) woodland in the Yalgorup area of south‐west Western Australia where the largest intact remaining example of this ecosystem is located. Although the cause of the decline is uncertain and several factors may be involved, management action should not be withheld because the decline process is not fully understood. We contend that the reduction in fire frequency over the last 50 years has led to an increase in understorey density, particularly of Western Australian Peppermint (Agonis flexuosa), resulting in greater competition for resources, which may in turn have increased the susceptibility of healthy woodland to decline. In contrast to Tuart regeneration, which is usually tied to fire, Western Australian Peppermint can establish readily in unburnt woodland. Further, once Western Australian Peppermint seedlings develop to the lignotuberous stage, they can resprout vigorously after fire. Therefore, a combination of fire and the physical removal of understorey in sites where this species has formed extensive thickets is required to: (i) provide an opportunity for regeneration of Tuart in both healthy and declining stands; (ii) improve the chances of sustained recovery of Tuart trees in declining stands; and (iii) ensure heterogeneity in the vegetation at multiple scales, a recognized strategy for conserving biodiversity and increasing ecosystem resilience. We propose that this approach may also be relevant to other tree decline syndromes in southern Australia. However, fostering community support for active intervention using thinning and fire in conservation reserves and staging the operations within an experimental framework will be important for such action to gain both the social and scientific acceptance necessary for it to be applied widely.