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Articles
Expression and purification of recombinant human serum albumin from selectively terminable transgenic rice
Qing Zhang, Hui Yu, Feng-zhen Zhang, Zhi-cheng Shen
Journal of Zhejiang University-SCIENCE B (Biomedicine & Biotechnology), 2013, 14(10): 867-874.   https://doi.org/10.1631/jzus.B1300090
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Human serum albumin (HSA) is widely utilized for medical purposes and biochemical research. Transgenic rice has proved to be an attractive bioreactor for mass production of recombinant HSA (rHSA). However, transgene spread is a major environmental and food safety concern for transgenic rice expressing proteins of medical value. This study aimed to develop a selectively terminable transgenic rice line expressing HSA in rice seeds, and a simple process for recovery and purification of rHSA for economical manufacture. An HSA expression cassette was inserted into a T-DNA vector encoding an RNA interference (RNAi) cassette suppressing the CYP81A6 gene. This gene detoxifies the herbicide bentazon and is linked to the 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) cassette which confers glyphosate tolerance. ANX Sepharose Fast Flow (ANX FF) anion exchange chromatography coupled with Butyl Sepharose High Performance (Butyl HP) hydrophobic interaction chromatography was used to purify rHSA. A transgenic rice line, HSA-84, was obtained with stable expression of rHSA of up to 0.72% of the total dry weight of the dehusked rice seeds. This line also demonstrated high sensitivity to bentazon, and thus could be killed selectively by a spray of bentazon. A two-step chromatography purification scheme was established to purify the rHSA from rice seeds to a purity of 99% with a recovery of 62.4%. Results from mass spectrometry and N-terminus sequencing suggested that the purified rHSA was identical to natural plasma-derived HSA. This study provides an alternative strategy for large-scale production of HSA with a built-in transgene safety control mechanism.
Genetic variability and evolution of rice stripe virus
Ling-zhe Huang, Li-xia Rao, Xue-ping Zhou, Jian-xiang Wu
Journal of Zhejiang University-SCIENCE B (Biomedicine & Biotechnology), 2013, 14(10): 875-885.   https://doi.org/10.1631/jzus.B1300014
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Rice stripe virus (RSV) is the type member of the genus Tenuivirus. RSV is known to have four segmented, single-stranded RNA molecules and causes rice stripe disease in the rice fields of China, Japan, and Korea. Based on the complete genomic sequences of the determined 6 RSV isolates (from Yunnan, Jiangsu, Zhejiang, and Liaoning Provinces, China) and 27 other RSV isolates (from Yunnan, Jiangsu, Anhui, Henan, and Shandong Provinces of China, also Japan and Korea) downloaded from GenBank, we provided a genotyping profile of RSV field isolates and described the population structure of RSV. All RSV isolates, except isolate CX, could be divided into two subtypes, one including 6 isolates from Yunnan Province, and the other including 26 isolates from different parts of China, Japan, and Korea, which were referred to as subtype II and subtype I, respectively. The amino acid distances between subtypes range from 0.053 to 0.085. RSV isolates in Yunnan Province were genetically differentiated from other parts of China, Japan, and Korea and showed infrequent gene flow. The RSV populations collected from other parts of China, Japan, and Korea were only composed of subtype I and showed very low genetic diversity. We speculated that isolate CX may be the result of recombination of isolates from two subtypes. Two potential recombination events were detected in RNA4 of isolate CX.
Quantitative changes of nitrergic neurons during postnatal development of chicken myenteric plexus
Ping Yang, Jameel Ahmed Gandahi, Qian Zhang, Lin-li Zhang, Xun-guang Bian, Li Wu, Yi Liu, Qiu-sheng Chen
Journal of Zhejiang University-SCIENCE B (Biomedicine & Biotechnology), 2013, 14(10): 886-895.   https://doi.org/10.1631/jzus.B1300005
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Objective: Information regarding the development of the enteric nervous system (ENS) is important for understanding the functional abnormalities of the gut. Because fertilized chicken eggs provide easy access to embryos, chicken models have been widely used to study embryonic development of myenteric plexus; however, no study has been focused on the postnatal period. The aim of this study was to perform a qualitative and quantitative analysis of the nitrergic neurons in the myenteric plexus of developing chickens in the postnatal period. Methods: Whole-mount preparations of the myenteric plexus were made in 7-d, 15-d, and 40-d old (adult) chickens of either sex (n=15). The myenteric plexus was studied after nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) histochemistry using light microscopy, digital photography, and Image-Pro Plus 6.0 software. The numbers of positively stained neurons and ganglia were counted in the duodenum, jejunum, ileum, caecum, and colon in the different age groups. Data were expressed as mean±standard deviation (SD), and statistical analysis was performed using a one-way analysis of variance (ANOVA) test. Results: The positively stained neurons showed various morphologies and staining intensities, and formed bead-shaped and U-shaped arrangements in the myenteric plexus. The densities of neurons and ganglia increased with age. However, the number of positive neurons per ganglion increased. The number of NADPH-d-positive neurons was highest in the colon, followed by the ileum, the jejunum, the duodenum, and the caeca in all age groups. Conclusions: Developmental changes in the myenteric plexus of chickens continue in the postnatal period, indicating that the maturation process of the gastrointestinal function is gradual. In addition, no significant difference is happening among different intestinal segments during postnatal development, suggesting that the function of different intestinal segments had been determined after birth.
Meat quality traits of four Chinese indigenous chicken breeds and one commercial broiler stock
Rong-fa Guan, Fei Lyu, Xiao-qiang Chen, Jie-qing Ma, Han Jiang, Chao-geng Xiao
Journal of Zhejiang University-SCIENCE B (Biomedicine & Biotechnology), 2013, 14(10): 896-902.   https://doi.org/10.1631/jzus.B1300163
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Meat quality traits of four genotypes of Chinese indigenous chicken [Ninghai chicken (NC), frizzle chicken (FC), Ninghai xiang chicken (XC), and Zhenning loquat chicken (LC)] and one genotype of commercial broiler [Arbor Acres plus broiler (AAB)] were analyzed. The indigenous chickens were raised before the commercial chickens in order to achieve the same final processed days. Indigenous chickens of NC, FC, XC, and LC showed significantly higher inosine-5′-monophosphate (IMP) content, shorter fiber diameter, and lower shear force than those of AAB (P<0.05). In the indigenous genotypes, NC and FC had significantly shorter fiber diameters and lower shear forces than XC and LC (P<0.05), and NC and XC had a higher IMP content than FC and LC (P<0.05). Moreover, the indigenous genotype of LC significantly displayed the highest protein content (P<0.05) in the five genotypes of birds, and no significant differences of protein content were found between the other genotypes of NC, FC, XC, and AAB (P>0.05). The indigenous chickens from FC displayed the highest total lipid content in the five bird genotypes (P<0.05). Significant differences of pH, color values of L* and a*, and drip loss for the five genotypes of birds were also observed. In conclusion, there were significant differences in the meat quality traits of the bird breeds selected in this study, and the indigenous chickens, especially the NC genotype, produced better quality meat as far as the IMP content, fiber diameters, and shear forces were concerned.
Optimization of extraction of phenolics from leaves of Ficus virens
Xiao-xin Chen, Xiao-bing Wu, Wei-ming Chai, Hui-ling Feng, Yan Shi, Han-tao Zhou, Qing-xi Chen
Journal of Zhejiang University-SCIENCE B (Biomedicine & Biotechnology), 2013, 14(10): 903-915.   https://doi.org/10.1631/jzus.B1200365
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In this research, the conditions for extraction of phenolics from leaves of Ficus virens were optimized using response surface methodology (RSM). The extraction abilities of phenolics (EAP) and flavonoids (EAF), the 2,2-diphenyl-1-pierylhydrazyl (DPPH) free-radical scavenging potential, and the ferric reducing/antioxidant power (FRAP) were used as quality indicators. The results of single-factor experiments showed that temperature, ethanol concentration, extraction time, and the number of extraction cycles were the main influencing variables, and these provided key information for the central composite design. The results of RSM fitted well to a second degree polynomial model and more than 98% of the variability was explained. The ideal extraction conditions for EAP, EAF, DPPH free-radical scavenging potential, and FRAP were obtained. Considering the four quality indicators overall, the ideal extraction conditions were 58% ethanol at 57 °C for 37 min with three extraction cycles. At the ideal extraction conditions, the values of EAP, EAF, DPPH free-radical scavenging potential, and FRAP were 5.72%, 3.09%, 58.88 mg ascorbic acid equivalent (AAE)/g dry weight (DW), and 15.86 mg AAE/g DW, respectively. In addition, linear correlations were observed between EAP, EAF, and antioxidant potential.
Identification of miRNAs and their targets in tea (Camellia sinensis)
Quan-wu Zhu, Yao-ping Luo
Journal of Zhejiang University-SCIENCE B (Biomedicine & Biotechnology), 2013, 14(10): 916-923.   https://doi.org/10.1631/jzus.B1300006
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MicroRNAs (miRNAs) are endogenous small RNAs playing a crucial role in plant growth and development, as well as stress responses. Among them, some are highly evolutionally conserved in the plant kingdom, this provide a powerful strategy for identifying miRNAs in a new species. Tea (Camellia sinensis) is one of the most important commercial beverage crops in the world, but only a limited number of miRNAs have been identified. In the present study, a total of 14 new C. sinensis miRNAs were identified by expressed sequence tag (EST) analysis from 47452 available C. sinensis ESTs. These miRNAs potentially target 51 mRNAs, which can act as transcription factors, and participate in stress response, transmembrane transport, and signal transduction. Analysis of gene ontology (GO), based on these targets, suggested that 37 biological processes were involved, such as oxidation-reduction process, stress response, and transport. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis inferred that the identified miRNAs took part in 13 metabolic networks. Our study will help further understanding of the essential roles of miRNAs in C. sinensis growth and development, and stress response.
A pilot scale trickling filter with pebble gravel as media and its performance to remove chemical oxygen demand from synthetic brewery wastewater
Haimanot Habte Lemji, Hartmut Eckst?dt
Journal of Zhejiang University-SCIENCE B (Biomedicine & Biotechnology), 2013, 14(10): 924-933.   https://doi.org/10.1631/jzus.B1300057
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Evaluating the performance of a biotrickling filter for the treatment of wastewaters produced by a company manufacturing beer was the aim of this study. A pilot scale trickling filter filled with gravel was used as the experimental biofilter. Pilot scale plant experiments were made to evaluate the performance of the trickling filter aerobic and anaerobic biofilm systems for removal of chemical oxygen demand (COD) and nutrients from synthetic brewery wastewater. Performance evaluation data of the trickling filter were generated under different experimental conditions. The trickling filter had an average efficiency of (86.81±6.95)% as the hydraulic loading rate increased from 4.0 to 6.4 m3/(m2∙d). Various COD concentrations were used to adjust organic loading rates from 1.5 to 4.5 kg COD/(m3∙d). An average COD removal efficiency of (85.10±6.40)% was achieved in all wastewater concentrations at a hydraulic loading of 6.4 m3/(m2∙d). The results lead to a design organic load of 1.5 kg COD/(m3∙d) to reach an effluent COD in the range of 50–120 mg/L. As can be concluded from the results of this study, organic substances in brewery wastewater can be handled in a cost-effective and environmentally friendly manner using the gravel-filled trickling filter.
Multi-year monitoring of paddy rice planting area in Northeast China using MODIS time series data
Jing-jing Shi, Jing-feng Huang, Feng Zhang
Journal of Zhejiang University-SCIENCE B (Biomedicine & Biotechnology), 2013, 14(10): 934-946.   https://doi.org/10.1631/jzus.B1200352
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The objective of this study was to investigate the tempo-spatial distribution of paddy rice in Northeast China using moderate resolution imaging spectroradiometer (MODIS) data. We developed an algorithm for detection and estimation of the transplanting and flooding periods of paddy rice with a combination of enhanced vegetation index (EVI) and land surface water index with a central wavelength at 2130 nm (LSWI2130). In two intensive sites in Northeast China, fine resolution satellite imagery was used to validate the performance of the algorithm at pixel and 3×3 pixel window levels, respectively. The commission and omission errors in both of the intensive sites were approximately less than 20%. Based on the algorithm, annual distribution of paddy rice in Northeast China from 2001 to 2009 was mapped and analyzed. The results demonstrated that the MODIS-derived area was highly correlated with published agricultural statistical data with a coefficient of determination (R2) value of 0.847. It also revealed a sharp decline in 2003, especially in the Sanjiang Plain located in the northeast of Heilongjiang Province, due to the oversupply and price decline of rice in 2002. These results suggest that the approaches are available for accurate and reliable monitoring of rice cultivated areas and variation on a large scale.
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