[Objective] In this study, the quantitive detection of PCV2 （porcine circovirus type 2） in vitro was achieved. We aimed to establish two kinds of TaqMan real-time PCR methods based on PCV20RF1 and ORF2 respectively ...[Objective] In this study, the quantitive detection of PCV2 （porcine circovirus type 2） in vitro was achieved. We aimed to establish two kinds of TaqMan real-time PCR methods based on PCV20RF1 and ORF2 respectively and compare them. [Method] According to the relatively＇conserved sequences of PCV20RF1 and ORF2 registered in GenBank, two pairs of specific primers and TaqMan probes were designed and synthesized. Then the recombinant plasmids containing the whole sequences of PCV20RF1 and ORF2 were constructed to draw the standard curves through optimizing the reaction system and conditions. And thus two kinds of TaqMan real-time PCR detection methods based on the whole sequences of ORF1 and ORF2 respectively were constructed for PCV2. [Result] For the two established standard curves, the Ct values showed a good linear relationship with the loga- rithms of copy numbers of templates （F2〉0.99）. The amplification efficiency ranged from 90% to 110%. The amplifications all had a good repeatability with variation coefficients within groups all less than 5%. Moreover, the amplifications all had a good specificity. When the sequences of porcine parvovirus （PPV）, porcine circovirus type 1 （PCV1）, swine pseudorabies virus （PRV）, porcine reproductive and respiratory syndrome virus （PRRSV） were used as templates, the target sequence was not amplified. The amplifications also had a high sensitivity. The ORF1 detection method could reach 1.0x10T copies/;ul, and the ORF2 detection method could reach 1.0×10^2 copies/μl. The two established real-time PCR detection methods were used to detect the 80 clinical samples respectively. The results showed the magnitudes of 72 amplified samples were basically consistent between the 2 detection methods, while the magnitudes of the other 8 amplified samples were inconsistent. Then the 8 samples were detected with SYBR Green I real-time PCR method established based on the sequence of PCV2-1ike factor P1 by Wen et aL The PCV2-1ike factor P1 was amplified in all the 8 samples, indicating the 8 samples were all infected with PCV2-1ike factor P1. [Conclusion] The ORFl-based detection method has a higher accuracy, and it can be used for the rapid detection of PCV2.展开更多
The family of mammalian chitinases includes members both with and without glycohydrolase enzymatic activity against chitin, a polymer of N-acetylglucosamine. Chitin is the structural component of fungi, crustaceans, i...The family of mammalian chitinases includes members both with and without glycohydrolase enzymatic activity against chitin, a polymer of N-acetylglucosamine. Chitin is the structural component of fungi, crustaceans, insects and parasitic nematodes, but is completely absent in mammals. Exposure to antigens containing chitin- or chitin-like structures sometimes induces strong T helper type-I responses in mammals, which may be associated with the induction of mammalian chitinases. Chitinase 3-like-1 （CHI3L1）, a member of the mammalian chitinase family, is induced specifically during the course of inflammation in such disorders as inflammatory bowel disease, hepatitis and asthma. In addition, CHI3L1 is expressed and secreted by several types of solid tumors including glioblastoma, colon cancer, breast cancer and malignant melanoma. Although the exact function of CHI3L1 in inflammation and cancer is still largely unknown, CHI3L1 plays a pivotal role in exacerbating the inflammatory processes and in promoting angiogenesis and remodeling of the extracellular matrix. CHI3L1 may be highly involved in the chronic engagement of inflammation which potentiates development of epithelial tumorigenesis presumably by activating the mitogen-activated protein kinase and the protein kinase B signaling pathways. Anti-CHI3L1 antibodies or pan-chitinase inhibitors may have the potential to suppress CHI3Ll-mediated chronic inflammation and the subsequent carcinogenic change in epithelial cells.展开更多
As an important agronomic trait, inclination of leaves is crucial Ior crop architecture and grain yields. 10 understand the molecular mechanism controlling rice leaf angles, one rice leaf inclination2 （1c2, three all...As an important agronomic trait, inclination of leaves is crucial Ior crop architecture and grain yields. 10 understand the molecular mechanism controlling rice leaf angles, one rice leaf inclination2 （1c2, three alleles） mutant was identified and functionally characterized. Compared to wild-type plants, lc2 mutants have enlarged leaf angles due to increased cell division in the adaxial epidermis of lamina joint. The LC2 gene was isolated through positional cloning, and encodes a vernalization insensitive 3-like protein. Complementary expression of LC2 reversed the enlarged leaf angles of lc2 plants, confirming its role in controlling leaf inclination. LC2 is mainly expressed in the lamina joint during leaf development, and particularly, is induced by the phytohormones abscisic acid, gibberellic acid, auxin, and brassinosteroids. LC2 is localized in the nucleus and defects of LC2 result in altered expression of cell division and hormone-responsive genes, indicating an important role of LC2 in regulating leaf inclination and mediating hormone effects.展开更多
基金Supported by National Natural Science Foundation of China(31302071)Special Fund for Agro-scientific Research in the Public Interest(201303046)+1 种基金Jiangsu Agricultural Science and Technology Innovation Fund(CX(14)2045)"333 High-level Personnel Training Project"of Jiangsu Province(BRA2012194)~~
文摘[Objective] In this study, the quantitive detection of PCV2 （porcine circovirus type 2） in vitro was achieved. We aimed to establish two kinds of TaqMan real-time PCR methods based on PCV20RF1 and ORF2 respectively and compare them. [Method] According to the relatively＇conserved sequences of PCV20RF1 and ORF2 registered in GenBank, two pairs of specific primers and TaqMan probes were designed and synthesized. Then the recombinant plasmids containing the whole sequences of PCV20RF1 and ORF2 were constructed to draw the standard curves through optimizing the reaction system and conditions. And thus two kinds of TaqMan real-time PCR detection methods based on the whole sequences of ORF1 and ORF2 respectively were constructed for PCV2. [Result] For the two established standard curves, the Ct values showed a good linear relationship with the loga- rithms of copy numbers of templates （F2〉0.99）. The amplification efficiency ranged from 90% to 110%. The amplifications all had a good repeatability with variation coefficients within groups all less than 5%. Moreover, the amplifications all had a good specificity. When the sequences of porcine parvovirus （PPV）, porcine circovirus type 1 （PCV1）, swine pseudorabies virus （PRV）, porcine reproductive and respiratory syndrome virus （PRRSV） were used as templates, the target sequence was not amplified. The amplifications also had a high sensitivity. The ORF1 detection method could reach 1.0x10T copies/;ul, and the ORF2 detection method could reach 1.0×10^2 copies/μl. The two established real-time PCR detection methods were used to detect the 80 clinical samples respectively. The results showed the magnitudes of 72 amplified samples were basically consistent between the 2 detection methods, while the magnitudes of the other 8 amplified samples were inconsistent. Then the 8 samples were detected with SYBR Green I real-time PCR method established based on the sequence of PCV2-1ike factor P1 by Wen et aL The PCV2-1ike factor P1 was amplified in all the 8 samples, indicating the 8 samples were all infected with PCV2-1ike factor P1. [Conclusion] The ORFl-based detection method has a higher accuracy, and it can be used for the rapid detection of PCV2.
基金supported by the National Natural Science Foundation of China(No.30871008 and 81370306Open Research Fund Program of Hubei Provincial Key Laboratory of Biological Targeting Therapy,China(No.2007B04)
文摘本文旨在建立转基因联合抑制心脏钙/钙调素依赖性蛋白激酶II(Ca2+/calmodulin-dependent protein kinase II,Ca MKII)及内向整流性钾电流(Ik1)小鼠模型,初步探讨该联合抑制在基础和应激状态下对小鼠心脏电生理、结构与功能重构的影响。将动物分为4组:野生组(WT),转基因Ca MKII抑制组(AC3-I),转基因Ik1抑制组(Kir2.1-AAA),转基因联合抑制组(AC3-I+Kir2.1-AAA)。记录各组动物在基础状态下及注射异丙肾上腺素后的心电图;同时分别对各组动物进行心脏超声检查,以了解心脏重构情况;在酶解法分离单个左室心肌细胞后,采用全细胞膜片钳法记录心肌细胞Ik1和瞬间外向钾电流(Ito)。实验结果显示:在基础状态下,心电图显示各组动物之间的心率、PR间期和QRS间期均无显著差别;注射异丙基肾上腺素后,上述心电图指标和小鼠发生室性心律失常的情况在各组间也无显著差别;4组小鼠间M型(Motion-mode)和二维超声检测的各项指标无显著差异;全细胞膜片钳实验结果表明,AC3-I组动物心肌细胞的Ik1电流密度最大(P<0.01),WT组的次之(P<0.01),其他两组较WT组显著降低(P<0.01);同时,AC3-I组Ito电流密度显著高于其他三组(P<0.01),而其他三组间无明显差异。上述结果提示:转基因联合抑制Ca MKII和Ik1可使Ca MKII抑制的小鼠Ik1下调,在基础状态下不影响心脏的结构和功能变化,在注射异丙基肾上腺素后心律失常的发生亦无明显差别。本结果为进一步探讨联合抑制Ca MKII与Ik1在各种心脏疾病状态下能否更有效地拮抗心肌的不利重构打下了基础。
基金Supported by NIH (DK64289,DK74454,DK43351 and DK80070)grants from the Eli and Edythe L.Broad Medical FoundationAmerican Gastroenterological Association Foundation for Digestive Health and Nutrition to Mizoguchi E
文摘The family of mammalian chitinases includes members both with and without glycohydrolase enzymatic activity against chitin, a polymer of N-acetylglucosamine. Chitin is the structural component of fungi, crustaceans, insects and parasitic nematodes, but is completely absent in mammals. Exposure to antigens containing chitin- or chitin-like structures sometimes induces strong T helper type-I responses in mammals, which may be associated with the induction of mammalian chitinases. Chitinase 3-like-1 （CHI3L1）, a member of the mammalian chitinase family, is induced specifically during the course of inflammation in such disorders as inflammatory bowel disease, hepatitis and asthma. In addition, CHI3L1 is expressed and secreted by several types of solid tumors including glioblastoma, colon cancer, breast cancer and malignant melanoma. Although the exact function of CHI3L1 in inflammation and cancer is still largely unknown, CHI3L1 plays a pivotal role in exacerbating the inflammatory processes and in promoting angiogenesis and remodeling of the extracellular matrix. CHI3L1 may be highly involved in the chronic engagement of inflammation which potentiates development of epithelial tumorigenesis presumably by activating the mitogen-activated protein kinase and the protein kinase B signaling pathways. Anti-CHI3L1 antibodies or pan-chitinase inhibitors may have the potential to suppress CHI3Ll-mediated chronic inflammation and the subsequent carcinogenic change in epithelial cells.
文摘As an important agronomic trait, inclination of leaves is crucial Ior crop architecture and grain yields. 10 understand the molecular mechanism controlling rice leaf angles, one rice leaf inclination2 （1c2, three alleles） mutant was identified and functionally characterized. Compared to wild-type plants, lc2 mutants have enlarged leaf angles due to increased cell division in the adaxial epidermis of lamina joint. The LC2 gene was isolated through positional cloning, and encodes a vernalization insensitive 3-like protein. Complementary expression of LC2 reversed the enlarged leaf angles of lc2 plants, confirming its role in controlling leaf inclination. LC2 is mainly expressed in the lamina joint during leaf development, and particularly, is induced by the phytohormones abscisic acid, gibberellic acid, auxin, and brassinosteroids. LC2 is localized in the nucleus and defects of LC2 result in altered expression of cell division and hormone-responsive genes, indicating an important role of LC2 in regulating leaf inclination and mediating hormone effects.
