To investigate the existence of the major outer membrane protein (MOMP) gene LipL32 in 15 dominant Chinese strains of 15 serogroups of Leptospira interrogans and 2 international strains of 2 serogroups of Leptospira b...To investigate the existence of the major outer membrane protein (MOMP) gene LipL32 in 15 dominant Chinese strains of 15 serogroups of Leptospira interrogans and 2 international strains of 2 serogroups of Leptospira biflexa, and to clone and construct the expression system as well as to identify the recombinant proteins, genomic DNAs from strains of leptospira were prepared by routine phenol-chloroform method, and the fragments of the LipL32 gene with the whole length from the strains were amplified with high fidelity PCR. The target amplification products were sequenced after T-A cloning, and the expression system for the genes were thereby constructed. Expression of the recombinant proteins was identified by using SDS-PAGE after induction with IPTG at different dosages. Western blot assays with rabbit antiserum against the whole cell of TR/PatocⅠ of Leptospira and immunized serum with rMOMPs were used to determine the immunoreactivity and immunogenicity of the recombinant proteins. Microscopic agglutination test was used to determine the cross- agglutination titres in rabbit sera immunized with rMOMPs, and the cell adherence model of Leptospira was used to examine the blocking effects of rabbit antisera against these rMOMPs. It was found that the LipL32 gene could be found in all the 17 strains of Leptospira mentioned above with two different genotypes, i.e. LipL32/1 and LipL32/2. Amounts of expressions of rMOMP1 and rMOMP2 after IPTG accounted for 40% and 10% of the total bacterial proteins respectively. Both rMOMP1 and rMOMP2 could combine with the rabbit antiserum against leptospiral TR/PatocⅠ, and could induce the production of agglutination antibodies to these 17 strains of Leptospira with 1∶2 to 1∶64 MAT titres. The rabbit anti-rMOMP1 and anti-MOMP2 antibodies at 1∶2 to 1∶16 dilutions could efficiently block adherence of Leptospira. It concludes that all the Leptospira tested in the present study possess LipL32/1 or LipL32/2 genes, and the constructed expression system can express the rMOMP1 and rMOMP2. These recombinant proteins are showed to have good immunogenicity and satisfactory immunoreactivity.展开更多
Precise responses to changes in light quality are crucial for plant growth and development.For example,hypocotyls of shade-avoiding plants typically elongate under shade conditions.Although this typical shade-avoidanc...Precise responses to changes in light quality are crucial for plant growth and development.For example,hypocotyls of shade-avoiding plants typically elongate under shade conditions.Although this typical shade-avoidance response(TSR)has been studied in Arabidopsis(Arabidopsis thaliana),the molecular mechanisms underlying shade tolerance are poorly understood.Here we report that B.napus(Brassica napus)seedlings exhibit dual shade responses.In addition to the TSR,B.napus seedlings also display an atypical shade response(ASR),with shorter hypocotyls upon perception of early-shade cues.Genome-wide selective sweep analysis indicated that ASR is associated with light and auxin signaling.Moreover,genetic studies demonstrated that phytochrome A(BnphyA)promotes ASR,whereas BnphyB inhibits it.During ASR,YUCCA8 expression is activated by early-shade cues,leading to increased auxin biosynthesis.This inhibits hypocotyl elongation,as young B.napus seedlings are highly sensitive to auxin.Notably,two non-canonical AUXIN/INDOLE-3-ACETIC ACID(Aux/IAA)repressor genes,BnIAA32 and BnIAA34,are expressed during this early stage.BnIAA32 and BnIAA34 inhibit hypocotyl elongation under shade conditions,and mutations in BnIAA32 and BnIAA34 suppress ASR.Collectively,our study demonstrates that the temporal expression of BnIAA32 and BnIAA34 determines the behavior of B.napus seedlings following shade-induced auxin biosynthesis.展开更多
文摘To investigate the existence of the major outer membrane protein (MOMP) gene LipL32 in 15 dominant Chinese strains of 15 serogroups of Leptospira interrogans and 2 international strains of 2 serogroups of Leptospira biflexa, and to clone and construct the expression system as well as to identify the recombinant proteins, genomic DNAs from strains of leptospira were prepared by routine phenol-chloroform method, and the fragments of the LipL32 gene with the whole length from the strains were amplified with high fidelity PCR. The target amplification products were sequenced after T-A cloning, and the expression system for the genes were thereby constructed. Expression of the recombinant proteins was identified by using SDS-PAGE after induction with IPTG at different dosages. Western blot assays with rabbit antiserum against the whole cell of TR/PatocⅠ of Leptospira and immunized serum with rMOMPs were used to determine the immunoreactivity and immunogenicity of the recombinant proteins. Microscopic agglutination test was used to determine the cross- agglutination titres in rabbit sera immunized with rMOMPs, and the cell adherence model of Leptospira was used to examine the blocking effects of rabbit antisera against these rMOMPs. It was found that the LipL32 gene could be found in all the 17 strains of Leptospira mentioned above with two different genotypes, i.e. LipL32/1 and LipL32/2. Amounts of expressions of rMOMP1 and rMOMP2 after IPTG accounted for 40% and 10% of the total bacterial proteins respectively. Both rMOMP1 and rMOMP2 could combine with the rabbit antiserum against leptospiral TR/PatocⅠ, and could induce the production of agglutination antibodies to these 17 strains of Leptospira with 1∶2 to 1∶64 MAT titres. The rabbit anti-rMOMP1 and anti-MOMP2 antibodies at 1∶2 to 1∶16 dilutions could efficiently block adherence of Leptospira. It concludes that all the Leptospira tested in the present study possess LipL32/1 or LipL32/2 genes, and the constructed expression system can express the rMOMP1 and rMOMP2. These recombinant proteins are showed to have good immunogenicity and satisfactory immunoreactivity.
基金supported by the Scientific Innovation 2030 Project(2022ZD0400801)the National Key R&D Program of China(2022YFD1200400)+1 种基金the National Natural Science Foundation of China(32100190)the National Natural Science Fund for Excellent Young Scientists Fund Program(Overseas).
文摘Precise responses to changes in light quality are crucial for plant growth and development.For example,hypocotyls of shade-avoiding plants typically elongate under shade conditions.Although this typical shade-avoidance response(TSR)has been studied in Arabidopsis(Arabidopsis thaliana),the molecular mechanisms underlying shade tolerance are poorly understood.Here we report that B.napus(Brassica napus)seedlings exhibit dual shade responses.In addition to the TSR,B.napus seedlings also display an atypical shade response(ASR),with shorter hypocotyls upon perception of early-shade cues.Genome-wide selective sweep analysis indicated that ASR is associated with light and auxin signaling.Moreover,genetic studies demonstrated that phytochrome A(BnphyA)promotes ASR,whereas BnphyB inhibits it.During ASR,YUCCA8 expression is activated by early-shade cues,leading to increased auxin biosynthesis.This inhibits hypocotyl elongation,as young B.napus seedlings are highly sensitive to auxin.Notably,two non-canonical AUXIN/INDOLE-3-ACETIC ACID(Aux/IAA)repressor genes,BnIAA32 and BnIAA34,are expressed during this early stage.BnIAA32 and BnIAA34 inhibit hypocotyl elongation under shade conditions,and mutations in BnIAA32 and BnIAA34 suppress ASR.Collectively,our study demonstrates that the temporal expression of BnIAA32 and BnIAA34 determines the behavior of B.napus seedlings following shade-induced auxin biosynthesis.
文摘目的:对RGC-32在大鼠主动脉损伤后的表达情况作初步研究,为血管术后再狭窄防治提供新思路。方法:选用雄性SD大鼠16只,大鼠随机分为两组。手术组:8只大鼠行球囊损伤大鼠主动脉术,术后28 d处死。对照组:8只大鼠,除不插入2F-Fogarty球囊导管进行损伤外,其他处理措施均与手术组相同,术后28 d处死。处死后收集大鼠主动脉术标本。分别进行下列检测:1血管形态学检测:切片HE染色;2SP染色免疫组织化学检测RGC-32表达,高倍光学显微镜下观察并计算阳性细胞百分比;3Real Time RT-PCR检测各组RGC-32在mRNA水平的表达情况。结果:手术组与正常对比,血管内膜面积增多,差异有统计学意义(P<0.05),手术组与对照组相比,RGC-32明显升高,且RGC-32平滑肌升高更明显,手术组RGC-32阳性表达及RGC-32mRNA水平明显高于对照组(P<0.05)。结论:大鼠主动脉损伤后RGC-32表达水平均明显升高,可为治疗新生内膜增生寻找新的靶点。
