Hepadnaviruses, including human hepatitis B virus （HBV）, replicate through reverse transcription of an RNA intermediate, the pregenomic RNA （pgRNA）. Despite this kinship to retroviruses, there are fundamental diff...Hepadnaviruses, including human hepatitis B virus （HBV）, replicate through reverse transcription of an RNA intermediate, the pregenomic RNA （pgRNA）. Despite this kinship to retroviruses, there are fundamental differences beyond the fact that hepadnavirions contain DNA instead of RNA. Most peculiar is the initiation of reverse transcription： it occurs by protein-priming, is strictly committed to using an RNA hairpin on the pgRNA, ε, as template, and depends on cellular chaperones; moreover, proper replication can apparently occur only in the specialized environment of intact nucleocapsids. This complexity has hampered an in-depth mechanistic understanding. The recent successful reconstitution in the test tube of active replication initiation complexes from purified components, for duck HBV （DHBV）, now allows for the analysis of the biochemistry of hepadnaviral replication at the molecular level. Here we review the current state of knowledge at all steps of the hepadnaviral genome replication cycle, with emphasis on new insights that turned up by the use of such cellfree systems. At this time, they can, unfortunately, not be complemented by three-dimensional structural information on the involved components. However, at least for the ~ RNA element such information is emerging, raising expectations that combining biophysics with biochemistry and genetics will soon provide a powerful integrated approach for solving the many outstanding questions. The ultimate, though most challenging goal, will be to visualize the hepadnaviral reverse transcriptase in the act of synthesizing DNA, which will also have strong implications for drug development.展开更多
Technology development has always been one of the forces driving breakthroughs in biomedical research. Since the time of Thomas Morgan, Drosophilists have, step by step, developed powerful genetic tools for manipulati...Technology development has always been one of the forces driving breakthroughs in biomedical research. Since the time of Thomas Morgan, Drosophilists have, step by step, developed powerful genetic tools for manipulating and functionally dissecting the Drosophila genome, but room for improving these technologies and developing new techniques is still large, especially today as biologists start to study systematically the functional genomics of different model organisms, including humans, in a high-throughput manner. Here, we report, for the first time in Drosophila, a rapid, easy, and highly specific method for modifying the Drosophila genome at a very high efficiency by means of an improved transcription activator-like effector nuclease （TALEN） strategy. We took advantage of the very recently developed ＂unit assembly＂ strategy to assemble two pairs of specific TALENs designed to modify the yellow gene （on the sex chromosome） and a novel autosomal gene. The mRNAs of TALENs were subsequently injected into Drosophila embryos. From 31.2% of the injected Fo fertile flies, we detected inheritable modification involving the yellow gene. The entire process from construction of specific TALENs to detection of inheritable modifications can be accomplished within one month. The potential applications of this TALEN-mediated genome modification method in Drosophila are discussed.展开更多
Big Personal Data is growing explosively. Consequently, an increasing number of internet users are drowning in a sea of data. Big Personal Data has enormous commercial value; it is a new kind of data asset. An urgent ...Big Personal Data is growing explosively. Consequently, an increasing number of internet users are drowning in a sea of data. Big Personal Data has enormous commercial value; it is a new kind of data asset. An urgent problem has thus arisen in the data market： How to price Big Personal Data fairly and reasonably. This paper proposes a pricing model for Big Personal Data based on tuple granularity, with the help of comparative analysis of existing data pricing models and strategies. This model is put forward to implement positive rating and reverse pricing for Big Personal Data by investigating data attributes that affect data value, and analyzing how the value of data tuples varies with information entropy, weight value, data reference index, cost, and other factors. The model can be adjusted dynamically according to these parameters. With increases in data scale, reductions in its cost,and improvements in its quality, Big Personal Data users can thereby obtain greater benefits.展开更多
AIM: To analyze the association of HCV-RNA with peripheral blood mononuclear cells (PBMC) and to answer the question whether HCV-RNA positivity in PBMC is due to viral replication. METHODS: HCV-RNA was monitored in se...AIM: To analyze the association of HCV-RNA with peripheral blood mononuclear cells (PBMC) and to answer the question whether HCV-RNA positivity in PBMC is due to viral replication. METHODS: HCV-RNA was monitored in serum and PBMC preparations from 15 patients with chronic HCV infection before, during and after an IFN-alpha therapy using a nested RT/PCR technique. In a second approach, PBMC from healthy donors were incubated in HCV positive plasma. RESULTS: In the IFN-alpha responding patients,HCV-RNA disappeared first from total RNA preparations of PBMC and then from serum. In contrast, in relapsing patients, HCV-RNA reappeared first in serum and then in PBMC. A quantitative analysis of the HCV-RNA concentration in serum was performed before and after transition from detectable to non detectable HCV-RNA in PBMC-RNA and vice versa. When HCV-RNA was detectable in PBMC preparations, the HCV concentration in serum was significantly higher than the serum HCV-RNA concentration when HCV-RNA in PBMC was not detectable. Furthermore, at no time during the observation period was HCV specific RNA observed in PBMC, if HCV-RNA in serum was under the detection limit. Incubation of PBMC from healthy donors with several dilutions of HCV positive plasma for two hours showed a concentration dependent PCR positivity for HCV-RNA in reisolated PBMC. CONCLUSION: The detectability of HCV-RNA in total RNA from PBMC seems to depend on the HCV concentration in serum. Contamination or passive adsorption by circulating virus could be the reason for detection of HCV-RNA in PBMC preparations of chronically infected patients.展开更多
AIM:To evaluate the effects of tenofovir disoproxil fumarate(TDF)use during late pregnancy to reduce hepatitis B virus(HBV)transmission in highly viremic mothers.METHODS:This retrospective study included 45 pregnant p...AIM:To evaluate the effects of tenofovir disoproxil fumarate(TDF)use during late pregnancy to reduce hepatitis B virus(HBV)transmission in highly viremic mothers.METHODS:This retrospective study included 45 pregnant patients with hepatitis B e antigen(+)chronic hepatitis B and HBV DNA levels>107copies/mL who received TDF 300 mg/d from week 18 to 27 of gestation(n=21).Untreated pregnant patients served as controls(n =24).All infants received 200 IU of hepatitis B immune globulin(HBIG)within 24 h postpartum and 20μg of recombinant HBV vaccine at 4,8,and 24 wk.Perinatal transmission rate was determined by hepatitis B surface antigen and HBV DNA results in infants at week 28.RESULTS:At week 28,none of the infants of TDFtreated mothers had immunoprophylaxis failure,whereas2(8.3%)of the infants of control mothers had immunoprophylaxis failure(P=0.022).There were no differences between the groups in terms of adverse events in mothers or congenital deformities,gestational age,height,or weight in infants.At postpartum week 28,significantly more TDF-treated mothers had levels of HBV DNA<250 copies/mL and normalized alanine aminotransferase compared with controls(62%vs none,P<0.001;82%vs 61%,P=0.012,respectively).CONCLUSION:TDF therapy during the second or third trimester reduced perinatal transmission rates of HBV and no adverse events were observed in mothers or infants.展开更多
文摘Hepadnaviruses, including human hepatitis B virus （HBV）, replicate through reverse transcription of an RNA intermediate, the pregenomic RNA （pgRNA）. Despite this kinship to retroviruses, there are fundamental differences beyond the fact that hepadnavirions contain DNA instead of RNA. Most peculiar is the initiation of reverse transcription： it occurs by protein-priming, is strictly committed to using an RNA hairpin on the pgRNA, ε, as template, and depends on cellular chaperones; moreover, proper replication can apparently occur only in the specialized environment of intact nucleocapsids. This complexity has hampered an in-depth mechanistic understanding. The recent successful reconstitution in the test tube of active replication initiation complexes from purified components, for duck HBV （DHBV）, now allows for the analysis of the biochemistry of hepadnaviral replication at the molecular level. Here we review the current state of knowledge at all steps of the hepadnaviral genome replication cycle, with emphasis on new insights that turned up by the use of such cellfree systems. At this time, they can, unfortunately, not be complemented by three-dimensional structural information on the involved components. However, at least for the ~ RNA element such information is emerging, raising expectations that combining biophysics with biochemistry and genetics will soon provide a powerful integrated approach for solving the many outstanding questions. The ultimate, though most challenging goal, will be to visualize the hepadnaviral reverse transcriptase in the act of synthesizing DNA, which will also have strong implications for drug development.
基金supported by the grants from the 973 Program(Nos.2009CB918702 and 2012CB945101)the NSFC(Nos.31071087 and 31100889)+1 种基金W.-M.D.is supported by NIH grant R01GM072562National Science Foundation of USA(IOS-1052333)
文摘Technology development has always been one of the forces driving breakthroughs in biomedical research. Since the time of Thomas Morgan, Drosophilists have, step by step, developed powerful genetic tools for manipulating and functionally dissecting the Drosophila genome, but room for improving these technologies and developing new techniques is still large, especially today as biologists start to study systematically the functional genomics of different model organisms, including humans, in a high-throughput manner. Here, we report, for the first time in Drosophila, a rapid, easy, and highly specific method for modifying the Drosophila genome at a very high efficiency by means of an improved transcription activator-like effector nuclease （TALEN） strategy. We took advantage of the very recently developed ＂unit assembly＂ strategy to assemble two pairs of specific TALENs designed to modify the yellow gene （on the sex chromosome） and a novel autosomal gene. The mRNAs of TALENs were subsequently injected into Drosophila embryos. From 31.2% of the injected Fo fertile flies, we detected inheritable modification involving the yellow gene. The entire process from construction of specific TALENs to detection of inheritable modifications can be accomplished within one month. The potential applications of this TALEN-mediated genome modification method in Drosophila are discussed.
基金supported in part by the National Natural Science Foundation of China (Nos. 61332001, 61272104, and 61472050)the Science and Technology Planning Project of Sichuan Province (Nos. 2014JY0257, 2015GZ0103, and 2014-HM01-00326SF)
文摘Big Personal Data is growing explosively. Consequently, an increasing number of internet users are drowning in a sea of data. Big Personal Data has enormous commercial value; it is a new kind of data asset. An urgent problem has thus arisen in the data market： How to price Big Personal Data fairly and reasonably. This paper proposes a pricing model for Big Personal Data based on tuple granularity, with the help of comparative analysis of existing data pricing models and strategies. This model is put forward to implement positive rating and reverse pricing for Big Personal Data by investigating data attributes that affect data value, and analyzing how the value of data tuples varies with information entropy, weight value, data reference index, cost, and other factors. The model can be adjusted dynamically according to these parameters. With increases in data scale, reductions in its cost,and improvements in its quality, Big Personal Data users can thereby obtain greater benefits.
基金Supported by a grant of DFG (SFB 402 Teilprojekt C1 (Mihm))by a grant of Hoffmann La Roche (Grenzach-Wyhden, Germany)Part of the data has been presented as poster at the 1999 EASL-meeting in Neaples
文摘AIM: To analyze the association of HCV-RNA with peripheral blood mononuclear cells (PBMC) and to answer the question whether HCV-RNA positivity in PBMC is due to viral replication. METHODS: HCV-RNA was monitored in serum and PBMC preparations from 15 patients with chronic HCV infection before, during and after an IFN-alpha therapy using a nested RT/PCR technique. In a second approach, PBMC from healthy donors were incubated in HCV positive plasma. RESULTS: In the IFN-alpha responding patients,HCV-RNA disappeared first from total RNA preparations of PBMC and then from serum. In contrast, in relapsing patients, HCV-RNA reappeared first in serum and then in PBMC. A quantitative analysis of the HCV-RNA concentration in serum was performed before and after transition from detectable to non detectable HCV-RNA in PBMC-RNA and vice versa. When HCV-RNA was detectable in PBMC preparations, the HCV concentration in serum was significantly higher than the serum HCV-RNA concentration when HCV-RNA in PBMC was not detectable. Furthermore, at no time during the observation period was HCV specific RNA observed in PBMC, if HCV-RNA in serum was under the detection limit. Incubation of PBMC from healthy donors with several dilutions of HCV positive plasma for two hours showed a concentration dependent PCR positivity for HCV-RNA in reisolated PBMC. CONCLUSION: The detectability of HCV-RNA in total RNA from PBMC seems to depend on the HCV concentration in serum. Contamination or passive adsorption by circulating virus could be the reason for detection of HCV-RNA in PBMC preparations of chronically infected patients.
文摘AIM:To evaluate the effects of tenofovir disoproxil fumarate(TDF)use during late pregnancy to reduce hepatitis B virus(HBV)transmission in highly viremic mothers.METHODS:This retrospective study included 45 pregnant patients with hepatitis B e antigen(+)chronic hepatitis B and HBV DNA levels>107copies/mL who received TDF 300 mg/d from week 18 to 27 of gestation(n=21).Untreated pregnant patients served as controls(n =24).All infants received 200 IU of hepatitis B immune globulin(HBIG)within 24 h postpartum and 20μg of recombinant HBV vaccine at 4,8,and 24 wk.Perinatal transmission rate was determined by hepatitis B surface antigen and HBV DNA results in infants at week 28.RESULTS:At week 28,none of the infants of TDFtreated mothers had immunoprophylaxis failure,whereas2(8.3%)of the infants of control mothers had immunoprophylaxis failure(P=0.022).There were no differences between the groups in terms of adverse events in mothers or congenital deformities,gestational age,height,or weight in infants.At postpartum week 28,significantly more TDF-treated mothers had levels of HBV DNA<250 copies/mL and normalized alanine aminotransferase compared with controls(62%vs none,P<0.001;82%vs 61%,P=0.012,respectively).CONCLUSION:TDF therapy during the second or third trimester reduced perinatal transmission rates of HBV and no adverse events were observed in mothers or infants.
