Abstract:[Objective] To obtain the spiroplasma resources and investigate the characteristics of the spiroplasmas on the surface of flowers in China. [Methods] The spiroplasma morphology was examined by dark-field microscope and transmission electron microscopy. The biological characteristics of the spiroplasmas were investigated by using conventional culture-dependent method and phylogenetic analysis based on 16S rRNA gene and ITS (16~23S rDNA intergenic space) sequence comparisons. Phylogenetic analysss was performed using the software package MEGA4.0 after multiple align-ment of sequence data by CLUSTAL X. [Results] Four spiroplasma isolates were obtained from three kinds of flower surface. Isolates CNR-1 and CNR-2 were from Brassica napus; CNA-1and CRW-1 from Rhododendron simsii and Oxalis corymbosa respectively. All isolates grew well in R-2 liquid medium and exhibited contractive movements. The colonies of all isolates were circinal and grain-like in solid medium. Through electron microscopy, all isolates exhibited helicity during their growth phase. All isolates could pass through a 0.22 μm filtrate membrane and resist to penicillin (2000 U/mL). They must grow in medium with serum. Glucose could be used as their carbon source instead of sucrose. The ability to hydrolyze arginine varied from different spiroplasmas but urea could not be hydrolyzed. The phylogenetic rela-tionships based on 16S rDNA supported CRW-1 was close to S. clarkii, and others were close to S. melliferum. The phy-logenetic relationship based on ITS sequence supported CRW-1 formed a separate clade, and others were also close to Spiroplasma melliferum. [Conclusion] The result indicated that spiroplasma isolate CRW-1 might be a new species and other three isolates were S. melliferum, but this need further support of serological test.

Abstract:[Objective] The purpose of this study was to characterize sigL mutant in Bacillus thuringiensis (Bt) HD-73, and to determine the function of sigL gene of Bt. [Methods] We studied the growth speed of the sigL mutant and complementary strain in different nutrient medium with different amino acids as nitrogen source respectively. lacZ gene was fused with the promoter of the acoR gene and bkdR gene, and two recombined genes were expressed in sigL mutant strain and HD-73 wild strain sequentially. [Results] sigL mutant could not grow on arginine, proline, valine, isoleucine, glutamine, phenylalanine, methionine and tryptophane as the sole nitrogen source separately. Activity of β-galactosidase in sigL mu-tant strain was much lower than it in wild-type strain and sequence analysis showed that the domain of AcoR and BkdR are similar to the conserved domain of SigL-dependent transcriptional activator in Bt. [Conclusion] The deletion of sigL gene maybe blocked some important metabolic pathways. AcoR and BkdR are σL-dependent transcriptional activators in Bacillus thuringiensis strains, probably the operons which were regulated by AcoR and BkdR were also controlled by σL respectively.

Abstract:[Objective] The aim was to investigate the regulator of phzA1B1C1D1E1F1(phzA1) operon in Pseudomoas aeruginosa. [Methods] Mutants affecting phzA1 expression in the presence of subinhibitory concentrations of tetracyclin were selected by screening a transposon mutagenesis library. The transposon insertion sites in the mutants were determined by arbitrary PCR and subsequent PCR product sequencing. [Results] We found two mutants affecting the phzA1 operon expression. The transposon insertion sites in these two mutants were at the promoter region of gene PA0487 encoding a probable molybdenum transport regulator. Some genes involved in quorum sensing were also down-regulated in the mutant. [Conclusion] PA0487 was involved in regulating the expression of phzA1, possibly through the quorum sensing system.

Abstract:[Objective] To study the component of ORPs (Oxysterol-binding proteins-related proteins) family in the rice fungus (Magnaporthe grisea), and the functions of MgORP1 gene with growing development of M. grisea, we constructed two MgORP1 deletion mutants and a complementation mutant. [Methods] All proteins of ORPs have a conserved OSBP-related ligand-binding domain (ORD), which was compared by BLASTP with the sequences of M. grisea genomic DNA published in M.grisea database. One of ORP gene, designated as MgORP1, was deleted by gene replacement and then the phenotypes of △MgORP1 were characterized. [Results] Six putative ORP family proteins have been found in M. grisea, and the growth rate of hyphal colony in complete medium and conidiation of △MgORP1 were lower, whereas the ability of conidia germination, appressoria formation and pathogenicity were normal. [Conclusion] MgORP1 gene of M.grisea was probably involved in colony growth and conidiation.

Abstract:[Objective] Transforming the specific insecticidal gene Bt cry3A into the dominant resident endogenetic bacteria in intestines of Apriona germari (Hope) larvae to construct transgenic bacteria that can colonize and express the insecticidal gene Bt cry3A perfectly in intestines of Apriona germari (Hope) larvae. [Method] We isolated and identified the dominant resident endogenetic bacteria by traditional methods and molecular method based of 16S rDNA analysis. Two Escherichia coli - Bacillus thuringiensis shuttle plasmid pHT305a and pHT7911 which contained specific insecticidal gene Bt cry3A were transformed into two resident endogenetic bacteria Brevibacillus brevis Ag12 and Bacillus thuringiensis Ag13 isolated from A. germari larvae intestines respectively by electro-transformation. [Results] Eighteen species of bacteria have isolated and identified from Apriona germari larvae intestines and two of them (Brevibacillus brevis Ag12 and Bacillus thuringiensis Ag13) were selected as starting bacteria to recieve the Bt cry3A. The 4 transgenic engineering strains Ag12-7911, Ag12-305a, Ag13-7911 and Ag13-305a were obtained successfully and validated by testing the plasmid stability in recombinants, transformants vegetal properties, crystal poisonous protein observation, expressional protein SDS-PAGE. The Bt cry3A gene had been transformed into Brevibacillus brevis and Bacillus thuringiensis. Both bioassay and examination of the engineering strains in intestines after feeding them to larvae showed that all these trans-formant strains (Brevibacillus brevis Ag12-305a, Bacillus thurigiensis Ag13-305a, Brevibacillus brevis Ag12-7911 and Bacillus thurigiensis Ag13-7911) could colonize and express 65 kDa protoxin in intestines of A. germari larvae and had insecticidal activity. [Conclusion] We obtained four transgenic bacteria that can colonize and express the target insecti-cide gene Bt cry3A in A. germari larvae. They may be developed as a new insecticide.

Abstract:[Objective] To clone and identify CnB gene from Setosphaeria turcica and to perform the primary bioinformatic analysis. [Methods] Using RACE method, we isolated the completed cDNA sequence and DNA sequence of CnB gene from S. turcica. Corresponding structure and functions were predicted by the bioinformatic software. [Results] The results showed that CnB gene had 4 exons and 3 introns and the largest open reading frame was 525 bp which encoded a protein of 174 amino acid residues. The predicted secondary structure composition for the protein contained about 59.77% alpha helixes, 8.62% beta turn, 6.32% extended strand and 25.29% random coil. The CnB gene of S. turcica had 4 “EF-hand” calcium-binding regions, and showed 90% homology with CnB gene of P. nodorum, B. cinerea and Neurospora crassa. [Conclusion] The CnB gene was successfully cloned from S. turcica for the first time, providing a good foundation for further research on its function.

Abstract:[Objective] Ginsenosides are important bioactive compounds in Ginseng. Sugar chains of ginsenosides are closely related to the bioactivity. Rare ginsenosides can be achieved by modifying sugar chains through biotransformation . Samples of Ginseng root soils, collected from Changbai Mountain, were used to screen active microorganism which can transform total ginsenosides and single ginsenoside, so as to obtain anti-tumor components. [Methods] The strains were isolated and screened on solid transfer medium and yield transfer process for active strains. Then the active strains were tested for their biotransformation properties by using different purified ginsenosides, including ginsenoside Rg3. The biotransformation products were separated and purified through column chromatography on silica gel (300mesh), and identified by spectral analysis and physical constants. Simultaneously, the active strain was identified based on morphological characteristics of colonies, conidium and conidiophore, as well as ITS-5.8S rDNA sequences. [Results] Total 68 fungal strains were isolated and 12 active strains showed positive activity on total ginsenosides. One strain, SYP2353, was found to have the strong activity on Rg3. [Conclusion] The active strain SYP2353 was identified as Myrothecium verrucaria, and the bio-transformation products of Rg3 were identified to be rare ginsenoside Rh2 and aglycon PPD.

Abstract:[Objective] To investigate construction and expression of novel staphylokinase (SAK) mutants with Arg-Gly-Asp (RGD) motif at its N-terminus, and to develop a novel thrombolytic agent with fibrinolytic activity and anti-platelet activity. [Methods] The fragments of SAK mutants were inserted into expression plasmid pBV220 and recombinant vectors were transformed into E. coli BL21, The SAK mutants could be expressed by rose of the culture temperature from 30°C to 42°C. After purified by a 3-step chromatography, their fibrinolytic and anti-platelet aggregation activity were analyzed. [Results] SDS-PAGE analysis indicated that the SAK mutants occupied 50% of the total proteins in E coli BL21. After purified by the 3-step chromatography, the purity of SAK mutants were more than 95%. The Specific fibrinolytic activities of SAK mutants purified were 10.8×104 and 11.0×104 HU·mg-1.Anti-platelet aggregation activity were 10.72% and 19.71%, which were significantly higher than that of wild-type SAK. Thus the SAK variants were successfully expressed, and purified. The high purity and bioactivities of staphylokinase variants lay a good basis for manufacture and clinical application.

Abstract:[Objective] Amorphophallus soft rot disease, caused by Erwinia carotovora, affects Amorphophallus industry development. We identified and characterized protein APn5 against Erwinia carotovora, isolated from Bacillus subtilis strain BSn5. [Methods] Protein APn5 was purified from BSn5 culture by ammonium sulfate precipitation with 30% rela-tive saturation and ultrafiltration. Inhibition activity was tested by agar well diffusion assay. Protein APn5 was treated at different temperatures, pH conditions and proteinase. The growth curve of BSn5 was examined; meanwhile, the inhibition activity of supernatant of BSn5 culture in different growth phase was tested. Amino acid sequence of protein APn5 was analyzed by matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF-MS) and quadru-pole-time-of-flight (Q-TOF). [Results] Protein APn5 showed a narrow inhibition spectrum, mainly strongly inhibiting the growth of a few plant pathogenic bacteria. Protein APn5 was sensitive to high temperature and alkaline pH, and partial sensitive to trypsin, proteinase K and pronase E. During strain BSn5 growth phase, the inhibition activity was unstable, which would gradually lose on stationary phase and disappeared finally. [Conclusion] On the basis of the difference on inhibition spectrum and characteristics, protein APn5 was suggested as a novel antimicrobial protein.

Abstract:[Objective] We studied the effect of the signal peptide sequence (SPS) on the expression of Pseudomonas pseudoalcaligenes insecticidal protein gene (ppip). [Methods] We obtained the core pseudomonas pseudoalcaligenes insecticidal protein gene (cppip, ppip without the UTR and SPS) by PCR and ligated it into pCAMBIA2301 to generate plant express vector pCPPIP, which was then transformed into tobacco to investigate the insecticidal activity of cppip expression products by locust bioassays. The Kanamycin resistance segregation ratio was determined by the germination rate of T0-generation seeds of the transgenic tobacco. Integration of ppip into genomic DNA was detected by PCR and confirmed by Southern blotting. [Results] The bioassay with the 2nd and 3rd instar larvae of Locusta orthoptera showed that the crude proteins extracted from cppip transformed plants caused an average mortality of 83.37%. In contrast, the protein extracts from ppip transformed plants caused a much lower mortality (15.65%). The growth of locust was highly inhibited by the expression products of cppip when compared with the locusts fed with the protein extracts from wild type tobacco or tobacco transformed with intact ppip gene. [Conclusion] The results indicated that the SPS might affect the insecticidal activity of ppip expressed in plants. The data of this study are helpful for cost-effective genetic engi-neering of plants with ppip gene.

Abstract:[Objective] Bacteria are able to adapt to temperatures far below their optimum growth temperatures, and a set of proteins (cold shock proteins, CSPs) are strongly induced in response to a rapid decrease in growth temperature. We studied the key functions in cryoprotection against damage caused by freezing of these proteins. [Methods] NcoI-HindIII CspC and CspD fragments were cloned respectively between NcoI and HindIII in pNZ8148, the recombinants plasmid were subsequently transformed by electroporation into Lactobacillus lactisNZ9000. Overproduction of CSPs was achieved by the addition of different concentrations of nisin to cultures and was analyzed by SDS-PAGE. In order to study the cryoprotection of CspC and CspD, the growth curves including the control strain and CSP-overproducing strains were developed. The number of colony-forming units was determined just before freezing and after four consecutive freeze-thaw operations. [Results] The 7 kDa cold-shock protein CspD and 6.2 kDa cold-shock protein CspC were identi-fied respectively. [Conclusion] The results indicate that CspC improves the recovery of cells and CspD increases the vi-ability after freezing (30~40 folds).

Abstract:[Objective] The aim of this study is to isolate novel and efficient polycyclic aromatic hydrocarbon (PAH) degarading bacteria from deep sea. [Methods] Bacteria in deep-sea water sample from Indian Ocean were enriched in the medium with crude oil as the carbon source. PAH-degrading bacteria were isolated and their degradation potential was tested by Gas Chromatography-Mass Spectrometry. PCR-primers were designed to detect the gene encoding the large subunit of aromatic-ring-hydroxylating dioxygenase. [Results] A PAHs-degrading bacterium, named H25 was obtained. Several PAHs including 2-methynaphthalene, 2, 6-dimethynaphthalene, phenanthrene and dibenzothiopheneand dibenzofuran could be used as carbon sources for growth by strain H25. Analysis of 16S rDNA sequence showed that it belonged to genus Novosphingobium with highest similarity (96%) with previously described bacteria. Two fragments of the di-oxygenase gene were obtained by PCR with size of about 700bp, which were closest to the counterpart of N. aromati-civorans DSM12444 with 99.6% and 91.0% similarities. Furthermore, two fragments named H25I (2.9 kb) and H25II(4.5kb) containing the upstream and downstream sequences were obtained by another set of primers. [Conclusion] Strain H25 was a novel PAH-degrading bacterium in deep sea environment, which might play a role in bioattenuation of PAH in oceanic environments and has potential in bioremediation of PAH contaminated environment.

Abstract:[Objective] To obtain pyrene-degrading microorganism strains, and to use them for bioremediation of polycyclic aromatic hydrocarbons-contaminated soil. [Methods] pyrene-degrading strains were isolated by agar plate subliming method. They were identified based on morphological observation, physiological and biochemical characteristics, and on analysis of their 16S rDNA gene sequence homology. Their ability to degrade PAHs in solid and liquid mineral salt medium, and in PAHs-contaminated soil, was studied by counting their live cells and measuring residual PAHs’ quantity by HPLC. [Results] Four pyrene-degrading strains TZh51, TZh52, TG42 and TG52 were isolated. The results indicated that TZh51’s ability to degrade PAHs was stronger than the other three strains. TZh51 was identified as Mycobacterium sp.; however, it did not belong to the same species with the reported Mycobacterium sp. strain M11. Our results indicated that the effects on maximal pyrene-degrading quantity were incubation temperature at 35℃and pyrene film thickness for 130ng/mm2 when TZh51 was incubated on solid mineral salt medium coated with pyrene films. When TZh51 was incu-bated in liquid mineral salt medium containing 50 and 100 mg/L pyrene, 91.9% and 71.8% pyrene was degraded after 6 days. On the10th day, maximal vital cells reached 2.0×108 and 6.0×108 cfu/mL, respectively. TZh51’s ability to degrade pyrene was stronger than strain M11. Moreover, TZh51’s vital cells reached 7.2×108 cfu/g dry soil on the 6th week and 91.4% phenanthrene, 86.9% fluoranthrene and 85.8% pyrene was degraded after 8 weeks, when both TZh51 and plants were used for bioremediation of PAHs-contaminated soil. [Conclusion] TZh51 had strong ability to degrade PAHs. In addition, combined bioremediation of TZh51 and plants was effective for PAHs-contaminated soil bioremediation.

Abstract:[Objective] We used Rhodopseudomonas strains with high-yield of 5-aminolevulinic acid (ALA) to produce ALA from wastewater of producing monosodium glutamate, citric acid, beer, and soybean product. [Methods] Cultivation was carried out under anaerobic light condition (3000 Lux) at 30℃. For comparison, we tested the addition of levulinic acid (LA), glycin and succinate to the substrate to increase the production of ALA, effect of sterilization of the wastewater for both strains. Cell mass concentration (OD660) and the content of ALA were determined with spectrophotometer. [Re-sults] Without adding levulinic acid (LA), glycin and succinate, the growth of strain 99-28 reached plateau after 72-96 h. The maxiam ALA production was obtained at 96 h. Both the yield of ALA and the Chemical Oxygen Demand (CODcr) removal rate of monosodium glutamate waster water were the highest in all tested wasterwaters. When LA, glycin and succinate were added, ALA production of strain 99-28 was significantly increased whereas the CODcr removal was ad-versely affected. Non-sterial wasterwater slightly reduced the growth and CODcr removal rate of strain 99-28, however the ALA production could be strongly reduced with the addition of LA, glycin and succinate. The growth and CODcr re-moval of mutant strain L-1 was similar with strain 99-28, but its ALA production was much higher than that of strain 99-28. [Conclusion] The Rhodopseudomonas strains screened in our laboratory can use organic wasterwater as substrates to produce ALA and remove CODcr.

Abstract:[Objective] To construct an stx2 gene mutant phage FMin27(Δstx∷cat) and to observe its infectiousness of various serotypes Escherichia coli strains. [Methods] With the help of Red recombinant system, the stx2 gene of the E. coli O157:H7 Min27 strain isolated from intestinal feces of piglet with diarrhea at a swine farm of Shanghai, was replaced by the chloramphenicol acetyltrasferase (cat) gene from plasmid pLacI. Phage FMin27(Δstx∷cat) was isolated after in-duction of E. coli Min27(Δstx∷cat) strain with mitomycin C. Twenty-one E. coli strains with various serotypes were infected with FMin27(Δstx∷cat), and plaque formation and lysogenic conversion of them were investigated. [Results] Of the 21 E. coli isolates, 2 with the serotypes of O60 and O138 integrated the FMin27(Δstx∷cat) in their chromosomes and expressed resistance to chloramphenicol. With the exception of one laboratory E. coli strain MG1655, none of the tested E. coli strains supported the formation of plaques and lysogenization when used as indicators for FMin27(Δstx∷cat). Fol-lowing induction with mitomycin C, these lysogenic strains released infectious particles of FMin27(Δstx∷cat) that formed plaques on a lawn of E.coli laboratory strain MC1061. [Conclusion] These results demonstrated that FMin27(Δstx∷cat) was able to infect and lysogenize particular E. coli strains and that the lysogens could produce infec-tious phage progeny. It could be inferred that Stx bacteriophages were able to spread exogenous genes among E. coli strains. The work provided a basis for further study on mechanisms of Stx phages infection and control of Stx expression.

Abstract:[Objective] To construct infectious DNA clone of chimeric porcine circovirus type 1-2. [Methods] Recombinant plasmid pSK-PCV1△ORF2-PCV2-ORF2 (pSK-sPCV1-2) was constructed by replacing the PCV1 capsid gene with that of PCV2 in the backbone recombinant containing the complete genomic DNA of PCV1. The full-length chimeric fragment PCV1-2 was excised from pSK-sPCV1-2 and cloned into the same recombinant to form the dimerized tandem DNA clone pSK-dPCV1-2. [Results] The sequence of PCV1-2 infectious clone was confirmed by sequencing. PCV1-2 virus was observed to be infectious after transfecting into both PK-15 and Dulac cells. BALB/c mice were immunized with recombinant virus PCV1-2 and sera samples collected from all control and vaccinated animals at -1, 7, 14, 21, 28, 35, 42 day post-inoculation (dpi) were assayed for anti-PCV2 capsid antibodies by ELISA. The results indicated that the chimeric PCV1-2 virus with the immunogenic ORF2 capsid gene of pathogenic PCV2 cloned into the nonpathogenic PCV1 genomic backbone induced specific antibodies against the pathogenic PCV2 capsid antigen in almost all mice at 42 dpi. [Conclusion] PCV1-2 infectious clone was constructed, and the recombinant virus strain of PCV1-2 was of some immunogenicity.

Abstract:[Objective] Avian influenza viruses are important human and animal respiratory pathogens and rapid diagnosis of novel emerging avian influenza viruses is vital for effective global influenza surveillance. We developed an oligonucleotide microarray-based method for subtyping all avian influenza virus (16 HA and 9 NA subtypes). [Methods] In total 25 pairs of primers specific for different subtypes and 1 pair of universal primers were carefully designed based on the genomic sequences of influenza A viruses retrieved from GenBank database. Several multiplex RT-PCR methods were then developed, and the target cDNAs of 25 subtype viruses were amplified by RT-PCR or overlapping PCR for evaluating the microarray. Further 52 oligonucleotide probes specific for all 25 subtype viruses were designed according to published gene sequences of avian influenza viruses in amplified target cDNAs domains, and a microarray for subtyping influenza A virus was developed. Then its specificity and sensitivity were validated by using different subtype strains and 2653 sam-ples from 49 different areas. [Results] The results showed that all the subtypes of influenza virus could be identified si-multaneously on this microarray with high sensitivity, which could reach to 2.47 pfu/mL virus or 2.5 ng target DNA. Fur-thermore, there was no cross reaction with other avian respiratory virus. [Conclusion] An oligonucleotide microar-ray-based strategy for detection of avian influenza viruses has been developed. Such a diagnostic microarray will be useful in discovering and identifying all subtypes of avian influenza virus.

Abstract:[Objective] To localize and identify the insecticidal crystal protein genes (cry genes) in high-toxic Bacillus thuringiensis strain 4.0718. [Methods] The genomic DNA of B. thuringiensis strain 4.0718 was isolated by pulse field gel electrophoresis (PFGE), and the PFGE pattern and plasmid pattern were characterized. The cry genes in the strain were analyzed by Southern blot and the cry gene types on the chromosome and plasmid were identified by PCR and restriction fragment length polymorphism patterns of the PCR-amplified fragments (PCR-RFLP). [Results] Southern blots showed that the cry genes in B. thuringiensis strain 4.0718 are present on both chromosome and plasmid. PCR and RFLP patterns showed that the chromosome and plasmid contain four cry genes, cry1Aa, cry1Ac, cry2Aa, and cry2Ab. However, the cry genes on the chromosome may be incomplete, while the cry1Aa and cry1Ac genes localized on the plasmid were found to contain the full coding sequence. [Conclusion] The abundant cry genes were first found on chromosome of B. thur-ingiensis strain 4.0718, and the cry-type genes are the same as the plasmid.苏云金芽胞杆菌；cry基因；定位；脉冲电泳（PFGE）；限制性片段长度多态性分析（RFLP）

Abstract:[Objective] Lactococcus lactis undergoing respiration has fast growth rate and relatively high biomass, which is the one of the potential way of improving the production efficiency of starter. In this study, the respiration situation and metabolism changes of Lactococcus lactis strains in the presence of heme were investigated. [Methods] Twelve strains of Lactococcus lactis were examined with aerobic cultivating test. The difference of biomass and metabolites among strains were compared. Furthermore, the survival rate at storage at 4℃ for 30 days was also compared by testing their viable cell count. [Results] A strain of Lactococcus lactis coded as KLDS 4.0316 could respire in the presence of heme. The biomass of KLDS 4.0316 cultivated in the present of heme increased by 50%. After being stored at 4℃ for 30 days, the strain cultivated under the condition without oxygen and heme was totally dead, whereas the viable cell count of strain cultivated with oxygen and heme remained 108 CFU/ml. Production of lactic acid decreased by 48%, compared with the strain cul-tivated in the absent of heme. [Conclusion] KLDS 4.0316 can undergo respiration in the present of heme, and the pro-duction of lactic acid decreased while the biomass increased.

Abstract:[Objective] The adenylation domain is required for the substrate activation of non-ribosomal peptide synthesis. The objective of this research was to prove that 2, 3-diaminopropionate is one of the presume precursors of Zwittermicin A biosynthesis. [Methods]We cloned the adenylation domain in the Zwittermicin A synthesis cluster of Bacillus thuringien-sis strain YBT-1520 with PCR amplification. After a series of enzyme digestions and subclonings, new expression vectors pBMB1312 was obtained. In order to detect the proper condition for overexpression, we tried different Isopropyl β-D-1-thiogalactopyranoside (IPTG) concentration and temperature during overepression. [Results]The overexpression protein of this domain could be purified under 20℃, 0.1 mmol/L Isopropyl b-D-1-thiogalactopyranoside (IPTG), BL21 codon plus RP（DE3）as the host strain. Then, PPi release assay indicated that 2, 3-diaminopropionate, the presume precursor of Zwittermicin A, could be adenylated by the adenylation domain. [Conclusion] This research confirmed that 2, 3-diaminopropionate is one of the presume precursors of Zwittermicin A biosynthesis.

Abstract:[Objective] To purify and characterize recombinant dextransucrase expressed in engineered strain BL21 (DE3)/pET28-dexYG. [Methods] The dextransucrase gene (dexYG) was expressed in engineered strain after IPTG induction and the crude enzyme was obtained by sonication. We purified the recombinant dextransucrase by using ammonium sulfate precipitation and metal chelate affinity chromatography on a Ni-NTA column. Then we characterized catalytic kinetic parameter of purified enzyme. [Results] This purification protocol resulted in a 11.4-fold purification with a yield of 37.5%. The molecular weight of dextransucrase measured by SDS-PAGE was 170 kDa ,which was similar to the enzyme from Leuconostoc mesenteroides. The enzyme had an optimum temperature between 25and 30℃ and an optimum pH of 5.4. It was relatively stable in the range of pH 5.0 to 7.0, but the stability declined rapidly as soon as the temperature rose over 35℃.The enzyme activity remained 59% after stored for 4 days at room temperature (25℃), and lost 50% activity after stored for seven weeks at 4℃. Ca2+ of 0.5 mmol/L could strongly activate the enzyme, Mg2+ of 1 mmol/L had little effect, Cu2+ and SDS could greatly inhibit the enzyme. [Conclusion] These results may provide an important basis for industrial applications of the recombinant dxtransucrase.

Abstract:With the advances and cost reducing of DNA-sequencing, genome determination for many microorganisms becomes available. For the research of lactic acid bacteria (LAB), this progress will not only facilitate to understand their nature of hereditary constitution, but also gradually leads to the possibility for analysis and further control of their biological functions. At present, the whole genome for 22 strains of LAB have been completed and published, at least another 12 strains of LAB are under sequencing. Based on the published data, we described the research progression of LAB ge-nomics from four aspects, including general features, diversity of metabolism, evolution and colinearity.