Abstract:Fresh fruit and vegetables are the basic components in people's daily diet, and play an important strategic role in the development of national economy. However, fresh fruit and vegetables are perishable, causing many problems during the logistics process, such as limited shelf life, high loss rate and serious quality deterioration, which are difficult to meet the rapid development of the industry and people's demand for high-quality life. Based on the perspective of dynamic logistics preservation of fresh fruit and vegetables, the research progress and frontier deveopment of domestic and foreign fresh fruit and vegetable logistics and packaging technology are comprehensively reviewed including post-harvest pre-cooling technology, logistics packaging materials and technology, cold storage and thermal insulation technology, quality monitoring technology during logistics process, and so on. The future prospect of research trend is also summarized, aiming to provide reference for scientific and technological innovation and industrial development of logistics packaging technology on fresh fruits and vegetables.

Abstract:Three novel polypeptides were designed with trypsin hydrolyzing residues for manufacturing anti-oxidant oligopeptides. These polypeptides successfully expressed in E. coli BL21(DE3) were named as ATO, ATW and WIT, respectively. Three kinds of polypeptide hydrolysates were achieved with trypsin digestion. In vitro antioxidant ability evaluation revealed that the generated polypeptide hydrolysates performed scavenging activity towards different free radicals. In comparison, the ATW hydrolysates showed the highest antioxidant activity, of which the IC50 values of DPPH free radical, superoxide anion free radical and hydroxyl free radical were 0.53 mg/mL, 0.45 mg/mL and 0.34 mg/mL, respectively. The present study provided a novel approach for antioxidant peptides design.

Abstract:In order to study the effects of different separation sources on the genome and function, 33 Lactobacillus plantarum from fermented sauce, kimchi and feces were selected. The basic characteristics of the genome, orthologous genes and system evolutionary relationshipwere studied by comparative genomics, combined with functional gene annotation results and phenotypic results to analyze strain resistance to antibiotics. The phylogenetic tree revealed a significant effect of the separation source on the genetic evolution of Lactobacillus plantarum. The homologous gene investigation showed that the fecal source strain had a higher specific gene number thanothers. 33 strains of Lactobacillus plantarum contain resistance genes of ciprofloxacin, tetracycline, chloramphenicol, trimethoprim and vancomycin, as well as their antibiotics. Most strains are sensitive to streptomycin, streptomycin, kanamycin, neomycinand ampicillin, and no related resistance genes were found in the genome. The genes and phenotypes of erythromycin and clindamycin do not correspond. The antibiotic experiments indicated an insignificant effect of the separation source on the strains and most of the genotypes and correspondence to the phenotype. Genomics is instructive to study the physiological characteristics of Lactobacillus plantarum.

Abstract:The preparation of melittin by biological method is a key technology for the large-scale preparation of melittin. To achieve efficient expression of melittin, the melittin gene met coding mature peptide and its promet containing the signal peptide, leader peptide and mature peptide were chemically synthesized. The two genes were linked with the maltose binding protein gene mbp and cloned into the vector pET15-b to construct the recombinant plasmid pET-MBP-MET and pET-MBP-proMET. The recombinant plasmids were then transformed into Escherichia coli BL21 (DE3) and induced with 0.1 mmol/L IPTG. The fusion protein MBP-MET and MBP-proMET were efficiently expressed in E. coli in soluble form treated overnight at 30 ℃. The two soluble target proteins were purified by two steps of Ni-affinity and Dextrin Sepharose HP affinity. The fusion proteins MBP-MET and MBP-proMET were obtained with an over 90% purity. About 20 mg of pure protein was prepared by 1 L of recombinant E. coli culture. The inhibition experiments of bacteria by MBP-MET and MBP-proMET were carried out. The obvious inhibition zone was observed in the Oxford Cup antibacterial experiments. The minimum inhibitory concentrations of MBP-MET and MBP-proMET were about 100 μg/mL and 140 μg/mL, respectively. Highly efficient expression was successfully achieved in this research. The function verification of melittin was confirmed, providing an efficient and safe synthetic route for the biological preparation of melittin.

Abstract:High-efficiency recombinant expression of Serratia marcescens non-specific nuclease (SMNE) was obtained using the prokaryotic Brevibacillus choshinensis expression system. The SMNE recombinant expression plasmid was constructed using the Escherichia coli-Bacillus spp. shuttle vector. After confirmation of the enzyme activity of SMNE in crude supernatant, the recombinant expression conditions were optimized towards temperature, glycerol content in the culture, and incubation time. The enzyme activity of recombinant SMNE wascharacterizedafter purificationby affinity and gel filtration chromatography. Recombinant SMNE was expressed in B. choshinensis HD31-SP3 strain. The initial enzyme activity in crude cell lysate was 4.07×10⁶ U/L. The expression condition was determined to be 5% glycerol at 30℃ for 56 h after optimization, which increased the activity to 2.6×107 U/L and obtained 6 folds of the initial activity yield. The purification procedure was also optimized and simplified to be a one-step affinity chromatography. The pure enzyme yield reached 30~40 mg/dL and the specific activity reached 1.3×10⁷ U/mg, which was 2~3 folds of that of the commercialcontrol. Characterization of the recombinant SMNE indicated that the optimal reaction condition was 37~45 ℃, pH 8~9. 47% of the activity remained without Mg²⁺/Mn²⁺, while 35%~45% of the activity remained with 300 mmol/L of Na⁺/K⁺.

Abstract:The protective effect of exogenous trehalose on brewer's yeast under heat stress was investigated via measuring the cell viability and vitality under 3 heat stress conditions, and the underlying mechanism was further studied via preliminary transcriptional analysis. As a result, exogenous trehalose was sufficient in improving the heat tolerance of brewer's yeast within the limited period and temperature ranges. The differences in the protective effects of various concentrations of trehaloseshrank with the increase of treatment temperature and duration. Moreover, the addition of exogenous trehalose elevated the intracellular trehalose level. Transcriptional analysis revealed that differentially expressed genes caused by trehaloseaddition mainly centered in ribosome biogenesis and related functions. It was thus speculated that the improved heat tolerance of brewer's yeast after adding exogenous trehalose to the culture medium was achieved by the elevated intracellular trehalose level. The increased level of intracellular trehalose promoted ribosome pathways andstrengthened the ribosome functions, further improving the heat tolerance of brewer's yeast.

Abstract:TAT-SOD is a fusion protein of TAT protein transduction domain and SOD. The TAT-SOD recombinant Pichia pastoris constructed in 2007 was used as the original strain to study the effects of different temperature,initial pH value, and inducer concentration on the expression of TAT-SOD in shake flask fermentation. Spectrophotometric analysis, scavenging of superoxide free radical and SDS-PAGE were used to measure the concentration of microorganism, the concentration of target protein TAT-SOD and the enzymatic activity of SOD, respectively. The mRNA changes of the target gene in the expressed strain were analyzed by quantitative PCR, aiming to investigate the suitability and optimization of the expression of TAT-SOD in long-term preserved recombinant Pichia pastoris strain. The results indicted that YPDM medium, inducer concentration of 1.0% (V/V), induction temperature of 30 ℃, initial pH value of 7.0 were the optimal condition of shake flask fermentation, with an enzymatic activity level in fermentation supernatant of 753 U/mL which was 5.1 times of that in the initial conditions．When the initial pH was 7.0, the expression level improved to 3.4 times and the mRNA level was 3.5 times of that under the control pH at 4.0. These results indicate that long-term preserved strain of recombinant Pichia pastoris is still suitable for the expression of TAT-SOD. The fermentation conditions significantly impact the expression of TAT-SOD, and the most important factor affecting the expression level is the mRNA level changes of the target protein.

Abstract:This research aimed to develop a neutral pullulanase with stable activity in a wide pH range, which can adapt to the manufacturing process of vermicelli with sweet potato powder as raw material. The pullulanse-encoding gene pul3665 was cloned from genome DNA of Geobacillus subterraneus XQ3665 by PCR and heterologously expressed. Specific activity of the purified recombinant enzyme PUL3665 is 32 U/mg. The optimal temperature and pH of PUL3665 were 55 ℃ and 6.5, respectively. The PUL3665 maintain 80% of its highest activity in the range of pH 5.5~7.7. The product of enzymatic degradation of pullulan was maltotriose, and the PUL3665 did not degrade amylose. Thus, the PUL3665 belongs to type I pullulanases. The crude enzyme of PUL3665 expressed by B. subtilis was used in the traditional vermicelli-making process. The paste was treated with PUL3665 at a dose of 2 U/g of paste. Vermicellis were successfully achieved using different kinds of sweet potato powder as raw materials. Neitheralum nor other additives were used. The quality of the finished vermicelli made from PUL3665 treated paste was about the same to that of the vermicelli which was made with alum as aditives. The PUL3665 is expectedto be developed into a highly effective and reliable substitute of alum in vermicelli production.

Abstract:In this study, the improvement of L-threonine synthesis in E. coli was achieved by optimizing transport of glucose. The CRISPR gene-editing method was used to delete the key genes ptsH and ptsG in the PTS system. The flask fermentation was carried out under 30 g/L glucose.Compared with the control strain TWF001, the productivity of L-threonine synthesis was significantly improved in either TWF001ΔptsH or TWF001ΔptsG, and the L-threonine yield of TWF001ΔptsH increased 38.02%. The strains TWF001ΔptsH, Ptrc::PgalP were successfully constructed via the enhancement of galP expression though adding the trc promoter in TWF001ΔptsH genome.Three mutant strains were grown by flask fermentation under 40, 50 or 60 g/L glucose, respectively. The fermentation yield of L-threonine in TWF001ΔptsH, Ptrc::PgalP after 36 h increased by 42.12%, which was 26.16 g/L with a sugar acid conversion rate of 0.65 g/g. The modification of glucose transport was confirmed to effectively improve L-threonine synthesis in E. coli.