Genome-Wide Identification of Hsp90 Gene Family in Perennial Ryegrass and Expression Analysis under Various Abiotic Stresses.

The heat shock protein 90 (Hsp90) is a protein produced in plants in response to stress. This study identified and analyzed Hsp90 gene family members in the perennial ryegrass genome. From the results, eight Hsp90 proteins were obtained and their MW, pI and number of amino acid bases varied. The amino acid bases ranged from 526 to 862. The CDS also ranged from 20 (LpHsp0-4) to 1 (LpHsp90-5). The least number of CDS regions was 1 (LpHsp90-5) with 528 kb amino acids, while the highest was 20 (LpHsp90-4) with 862 kb amino acids, which showed diversity among the protein sequences. The phylogenetic tree revealed that Hsp90 genes in Lolium perenne, Arabidopsis thaliana, Oryza sativa and Brachypodium distachyon could be divided into two groups with five paralogous gene pairs and three orthologous gene pairs. The expression analysis after perennial ryegrass was subjected to heat, salt, chromium (Cr), cadmium (Cd), polyethylene glycol (PEG) and abscisic acid (ABA) revealed that LpHsp90 genes were generally highly expressed under heat stress, but only two LpHsp90 proteins were expressed under Cr stresses. Additionally, the expression of the LpHsp90 proteins differed at each time point in all treatments. This study provides the basis for an understanding of the functions of LpHsp90 proteins in abiotic stress studies and in plant breeding.

Effect of naphthalene acetic acid on adventitious root development and associated physiological changes in stem cutting of Hemarthria compressa.

In order to find a way to induce rooting on cuttings of Hemarthria compressa cv. Ya'an under controlled conditions, a project was carried out to study the effect of naphthalene acetic acid (NAA) on rooting in stem cuttings and related physiological changes during the rooting process of Hemarthria compressa. The cuttings were treated with five concentrations of NAA (0, 100, 200 300, 400 mg/l) at three soaking durations (10, 20, 30 minutes), and cuttings without treatment were considered as control. Samples were planted immediately into pots after treatment. IAA-oxidase (IAAO) activity, peroxidase (POD) activity and polyphenol oxidase (PPO) activity were determined after planting. Results showed that NAA had positive effect on rooting at the concentration of 200 mg/l compared to other concentrations at 30 days after planting (DAP). Among the three soaking durations, 20 minutes (min) of 200 mg/l NAA resulted in higher percentages of rooting, larger numbers of adventitious roots and heavier root dry weight per cutting. The lowest IAAO activity was obtained when soaked at 200 mg/l NAA for 20 min soaking duration. This was consistent with the best rooting ability, indicating that the lower IAAO activity, the higher POD activity and PPO activity could be used as an indicator of better rooting ability for whip grass cuttings and might serve as a good marker for rooting ability in cuttings.

Purification and preliminary crystallographic studies of CutC, a novel copper homeostasis protein from Shigella flexneri.

CutC is a novel copper homeostasis protein containing 248 amino acids. Here we report the cloning, expression, purification, crystallization and preliminary X-ray crystallographic studies of CutC from Shigella flexneri 2a. Purification of CutC and its selenomethionine (SeMet) derivate were done using immobilized metal ion affinity chromatography, size-exclusion and ion-exchange chromatography. The purified proteins were crystallized using the hanging drop vapor diffusion method. The diffraction data for the native and SeMet CutC, respectively, have been collected with resolution of 1.7 A and 2.1 A. They belong to the space group C2221 and similar cell dimension. The native protein crystals have cell parameters: a=75.3267, b=97.6718, c=132.6910.

Optimizing protein crystal growth through dynamic seeding.

A dynamic seeding method that is different from the conventional method of seeding drops that have been equilibrated is described. The method basically consists of two steps. Firstly, microseeding was used in association with adjustment of the seeding-drop components, including buffer, additive and concentrations of the precipitants and protein, in order to screen suitable seeding conditions under which microseeds are seeded into a new non-equilibrated drop as the dynamic macroseed drop for the following step. Secondly, after being equilibrated for various times against the reservoir solution, the macroseed drops were used to prepare a dilution series with which the qualified crystals could be harvested using macroseeding. Compared with a conventional seeding technique, this method is distinct with a dynamic situation of macroseed drops before macroseeding and a non-equilibrium serial seeding where all the seeds are seeded into new non-equilibrated drops and the micro/macroseeding are efficiently combined into a whole system. The method simplifies control of the number of microseeds because an excess of microseeds has little effect on the final result. The method also simplifies the manipulation of macroseeds by optimizing the equilibration time and the dilution multiple of the macroseed drops before macroseeding. This dynamic seeding technique has been used in the crystallization of novel protein CutCm, which has a fast crystal-growth rate, and proved that the method is useful for optimizing protein crystallization.