Single-copy Loss of Rho Guanine Nucleotide Exchange Factor 10 ( arhgef10) Causes Locomotor Abnormalities in Zebrafish Larvae / 生物医学与环境科学（英文）

OBJECTIVE@#To determine if ARHGEF10 has a haploinsufficient effect and provide evidence to evaluate the severity, if any, during prenatal consultation.@*METHODS@#Zebrafish was used as a model for generating mutant. The pattern of arhgef10 expression in the early stages of zebrafish development was observed using whole-mount in situ hybridization (WISH). CRISPR/Cas9 was applied to generate a zebrafish model with a single-copy or homozygous arhgef10 deletion. Activity and light/dark tests were performed in arhgef10 -/-, arhgef10 +/-, and wild-type zebrafish larvae. ARHGEF10 was knocked down using small interferon RNA (siRNA) in the SH-SY5Y cell line, and cell proliferation and apoptosis were determined using the CCK-8 assay and Annexin V/PI staining, respectively.@*RESULTS@#WISH showed that during zebrafish embryonic development arhgef10 was expressed in the midbrain and hindbrain at 36-72 h post-fertilization (hpf) and in the hemopoietic system at 36-48 hpf. The zebrafish larvae with single-copy and homozygous arhgef10 deletions had lower exercise capacity and poorer responses to environmental changes compared to wild-type zebrafish larvae. Moreover, arhgef10 -/- zebrafish had more severe symptoms than arhgef10 +/- zebrafish. Knockdown of ARHGEF10 in human neuroblastoma cells led to decreased cell proliferation and increased cell apoptosis.@*CONCLUSION@#Based on our findings, ARHGEF10 appeared to have a haploinsufficiency effect.

Genetic analysis of an individual with a fragile site at 16q22 / 中华医学遗传学杂志

OBJECTIVE@#To analyze a patient with infertility and a fragile site found at 16q22 by using cytogenetic methods.@*METHODS@#Peripheral blood sample was taken from the patient and subjected to chromosomal karyotyping and single nucleotide polymorphism microarray (SNP-array) analysis.@*RESULTS@#The patient was found to be a mosaicism for a fragile site at 16q22, which has a variable morphology and cannot be induced by folic acid treatment. No abnormality was found by SNP-array analysis.@*CONCLUSION@#A rare fragile site, which can be induced without folic acid treatment, has been identified at 16q22. The strategy of assisted reproduction for such individuals is yet to be explored.

Influencing factors and pregnancy outcomes of unsuccessful cell-free DNA testing in maternal perinatal blood / 中华围产医学杂志

Objective:To explore the possible factors leading to failure of cell-free DNA (cfDNA) testing in maternal peripheral blood and analyze the pregnancy outcomes of this group of pregnant women.Methods:This retrospective study involved 5 195 women who underwent cfDNA testing in Peking University Third Hospital from April 2017 to April 2019. Based on the first cfDNA testing results, clinical characteristics of the pregnant women with successful (success group, n=5 107) and failed (failure group, n=88) cfDNA testing were compared using Mann-Whitney U test and Chi-square test. Multivariate logistic regression was used to analyze the risk factors of cfDNA testing failure and the effect of body mass index (BMI) on the success rate, and evaluate the feasibility of re-sampling and the factors affecting the unsuccessful testing of a second sample. Results:The failure rate of first cfDNA testing was 1.7% (88/5 195). Successful cfDNA testing was achieved in 74 (87.1%, 74/85) of 85 re-sampling cases, while results of the other 11 cases (12.9%, 11/85) remained invalid. Thus, the final failure rate was 0.2% (11/5 195). Multivariate logistic regression revealed that increased maternal age ( OR=1.086, 95% CI: 1.023-1.152, P=0.006), BMI ( OR=1.083, 95% CI: 1.021-1.149, P=0.008) and twin pregnancies ( OR=3.093, 95% CI: 1.715-5.577, P<0.001) were the risk factors of cfDNA testing failure, while increased cell-free fetal DNA (cffDNA) concentration ( OR=0.758, 95% CI: 0.720-0.761, P<0.001) was a protective factor. The overweight (BMI: 25-29.9 kg/m 2) and obese (BMI≥30 kg/m 2) women were 3.626 ( OR=3.626, 95% CI: 2.298-5.724, P<0.001) and 4.064 ( OR=4.064, 95% CI: 1.779-9.284, P=0.001) times more likely to have failed cfDNA testing than those with normal weight (BMI: 18.5-24.9 kg/m 2), respectively. The success rate of re-testing decreased as the maternal BMI increased, regardless of the time interval between the two samplings ( OR=0.840, 95% CI: 0.699-1.245, P=0.065). Seven out of the 74 cases with successful results in re-testing were at high risk, including one 45,X and one 47,XXY, confirmed by karyotyping amniocentesis. Among the 11 pregnant women with a failed testing after second sampling, eight underwent prenatal diagnosis with normal fetal chromosome karyotypes, and the other three cases without prenatal diagnosis all gave birth to neonates with normal phenotype. There was no statistical difference in the incidence of pregnancy loss between the failure and success group [9.1% (8/88) vs 2.5% (128/5 107), P=0.090]. Conclusions:Pregnant women with advanced age and higher BMI, lower cffDNA fraction and twin pregnancies are more likely to fail in cfDNA testing. For obese women, blood sampling can be postponed to a larger gestational age to reduce the failure rate. For pregnant women with failed testing in first sampling, a re-sampling is recommended, moreover, prenatal diagnosis is necessary for those had high-risk results or failed in re-testing.

Analysis of clinical factors on early pregnancy loss during in vitro fertilization-embryo transfer treatment and natural pregnancy / 中华妇产科杂志

Objective To compare differences of clinical factors related to early pregnancy loss between invitro fertilization-embryo transfer (IVF-ET) treatment and natural pegnancy. Methods A retrospective analysis was performed on the 363 cases of early pregnancy loss between Dec. 2015 to May 2016 in Peking University Third Hospital, during which 173 cases were after IVF-ET treatment(IVF-ET group), and others were natural pregnancies(natural group). Results The average age in IVF-ET group was significantly higher than that in the natural group [(34.1±4.3)versus(31.8±4.1)years old, P<0.01]. The terminating time of pregnancy loss in IVF-ET group was short than that in the natural group [(59.8±9.2) versus(69.9 ± 11.1)days, P<0.01]. The incidence of embryo abnormal chromosome in IVF-ET group was significantly lower than that in the natural group [57.2%(99/173)versus 74.2%(141/190), P<0.01], during which abnormal chromosome numbers were the most common. Conclusions The pregnancy loss of early pregnancy is mainly caused by chromosome abnormality. The proportion of chromosome abnormality in early pregnancy loss after IVF-ET is not higher than that of natural pregnancy, indicating that there are relatively reliable gametes and embryo safety in IVF treatment.

Bioinformatics analysis raises candidate genes in blood for early screening of Parkinson's disease / 生物医学与环境科学（英文）

Parkinson's disease (PD) is a typical degenerative disease, which is characterized by the most obvious symptoms of movement dysfunction, including shaking, rigidity, slowness of movement and difficulty in walking and gait. This disease can not be clearly identified through laboratory tests at present, thus application of high-throughput technique in studying the expression profiles of PD helps to find the genetic markers for its early diagnosis. Studies on expression profiles of neurodegenerative diseases have revealed the novel genes and pathways involved in the progress of illness. In this study, the expression profiles of PD in blood were compared, showing that 181 differentially expressed genes (DEG) exhibit a similar expression trend both in patients and in normal controls. These genes are enriched significantly in some biological processes, including development, response to drugs, and DNA-dependent regulation of transcription, etc, highlighting that the genetic markers can be used in early diagnosis of PD.

Clinical and familial characteristics of ten chinese patients with fatal family insomnia / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>Fatal familial insomnia (FFI) is an autosomal dominant prion disease characterized clinically by inattention, sleep loss, dysautonomia, and motor signs. This study is aimed to investigate clinical and familial characteristics of ten Chinese Patients with FFI.</p><p><b>METHODS</b>We identified ten FFI cases from the surveillance network for Creutafeldt-Jakob disease (CJD) in China. Final diagnosis of FFI cases was made in accordance with the WHO criteria for CJD. The main clinical features and family histories of these ten FFI cases were analyzed.</p><p><b>RESULTS</b>The median age of ten cases at onset was 38 years (from 19 to 55). The foremost symptoms seemed to be various, including sleep disturbances, vision disorder, dizziness and anorexia. Sleep disturbances appeared in all cases and lasted in the whole clinical courses. Progressive sympathetic symptoms, memory loss, movement disturbances, myoclonus and hypertension were also frequently observed. The median duration of the disease was 9.5 months. EEG and MRI did not figure out special abnormality. 14-3-3 protein in CSF was positive in five out of eight tested patients. Clear family histories were identified in 8 patients.</p><p><b>CONCLUSION</b>The data from our study confirm that the Chinese FFI cases have similar clinical characteristics as that of the Caucasian cases. Compared with other genetic CJD associated mutations, the genetic frequencies of D178N in PRNP are apparently high among the Chinese cases.</p>

Mutations in various functional domains of HPV2 E2 protein inhibit the transcriptional depression activities / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To study the potential transcriptional depression activities of HPV2 E2 proteins with mutations in different functional domains.</p><p><b>METHODS</b>The primers for constructing various E2 mutants were synthesized based on a HPV2 isolate containing several point mutations within E2 open reading frame. Different E2 mutations were generated by the method of extending PCR and inserted into plasmid pcDNA3. 1. Various recombinant mammalian expression plasmids pcDNA3. 1-E2 were co-transfected into HeLa cells together with a CAT-reporter plasmid pBLCAT-LCR containing HPV-2 prototype LCR, respectively. The transcriptional repression activities of the E2 mutants were evaluated by detection of CAT expression values.</p><p><b>RESULTS</b>Compared with the full-length prototype E2, removals of both N- and C-terminal domains abolished E2 transcriptional repressive activities. The point mutations in the transactivation domain (nt 3037), the internal hinge region (nt 3387) and DNA binding domain (nt 3697) showed remarkable inhibition on its transcriptional depression function.</p><p><b>CONCLUSION</b>The transcriptional regulation activity of HPV2 E2 is related with its DNA binding and transactivation domains. The exchanges of the single amino acid within E2, derived from a HPV2 isolate, abolish significantly the repressive effect on viral promoter in the context of full-length E2.</p>

Establishment of hamster- and human-PRNP transgenic mice / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To create transgenic mice expressing hamster- and human-PRNP as a model for understanding the physiological function and pathology of prion protein (PrP), as well as the mechanism of cross-species transmission of transmissible spongiform encephalopathies (TSEs).</p><p><b>METHODS</b>Hamster and human-PRNP transgenic mice were established by conventional methods. The copy number of integrated PRNP in various mouse lines was mapped by real-time PCR. PRNP mRNA and protein levels were determined by semi-quantitative RT-PCR, real-time RT-PCR, and western blot analysis. Histological analyses of transgenic mice were performed by hematoxylin and eosin (H & E) staining and immunohistochemical (IHC) methods.</p><p><b>RESULTS</b>Integrated PRNP copy number in various mouse lines was 53 (Tg-haPrP1), 18 (Tg-huPrP1), 3 (Tg-huPrP2), and 16 (Tg-huPrP5), respectively. Exogenous PrPs were expressed at both the transcriptional and translational level. Histological assays did not detect any abnormalities in brain or other organs.</p><p><b>CONCLUSION</b>We have established one hamster-PRNP transgenic mouse line and three human-PRNP transgenic mouse lines. These four transgenic mouse lines provide ideal models for additional research.</p>

Interaction between various 14-3-3beta segments and PrP in vitro / 中华实验和临床病毒学杂志

<p><b>UNLABELLED</b>OBJECTIVE To study the potential interaction between PrP protein.</p><p><b>METHODS</b>The supernatant of health and scrapie-infected hamsters' brain homogenate was prepared, while various recombinant 14-3-3beta or PrP proteins were purified. The possible molecular interaction between 14-3-3beta proteins and PrP was tested by pull-down and immunoprecipitation assays.</p><p><b>RESULTS</b>Both native PrP(c) and its protease-resistant isoform (PrP(Sc)) formed complexes with 14-3-3beta. The full-length recombinant 14-3-3beta proteins interacted with PrP. The domain responsible for interacting 14-3-3beta was located at N-terminal of 14-3-3beta (residues 1 to 38).</p><p><b>CONCLUSION</b>The studies of the association of PrP with 14-3-3beta may further provide insight into a potential role of 14-3-3beta in the biological function of PrP and the pathogenesis of prion disease.</p>

The first Chinese case of Creutzfeldt-Jakob disease with mutation of E200K in PRNP / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To investigate epidemiological, clinical and genetic features of the first Chinese case of Creutzfeldt-Jakob disease (CJD ) with mutation of E200K in PRNP.</p><p><b>METHODS</b>The general epidemiological and clinical data were collected; CSF 14-3-3 protein was analyzed by Western blot; The PRNP was amplified by PCR and analyzed.</p><p><b>RESULTS</b>A missense mutation in codon 200 (E200K) of the PRNP was identified in this patient; CSF 14-3-3 protein was positive; sleep disturbance was the initial sign and the other symptoms gradually appeared, including memory loss, dizziness and ataxia.</p><p><b>CONCLUSION</b>The CJD patient who was first reported in China has a missense mutation in codon 200 (E200K) of the PRNP, and the codon 129 is a methionine homozygous genotype.</p>

Cytotoxicity of human doppel protein and Dpl-like protein PrPΔ32-121 to SH-SY5Y cells / 西安交通大学学报(医学版)

Objective To observe the biological activities of human doppel (Dpl) protein transiently expressed and Dpl-like protein PrPΔ32-121 on a human neuroblastoma cell line SH-SY5Y. Methods Recombinant mammalian expression plasmids containing human PRND gene and truncated PrPΔ32-121 fragment were generated by PCR. The expression and location of Dpl and PrPΔ32-121 post-transfection were observed by IFA. The cytotoxicity was measured by MTT analysis. Cellular apoptosis was investigated by flow cytometry and Western blot. Results Both Dpl and PrPΔ32-121 protein were expressed and mainly located on the cell membrane. Remarkable cytotoxicity was detected on SH-SY5Y cells after 24 h transfection. Meanwhile, more Annexin V/PI positively-stained cells as well as lower levels of cellular pro-caspase-3 and Bel-2 were detected in the cells receiving Dpl and PrPΔ32-121 expressing plasmids. Conclusion Dpl protein transiently expressed and PrPΔ32-121 can lead to the similar neural cytotoxicity, probably triggering the cell apoptosis program.

Study on the phenomenon of splashes and sprays from virology / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To investigate the phenomenon of accidental splashes and sprays from manipulation of recombinant virus material and to measure the approximate spilled distance when recombinant virus material inadvertently dropped in the biosafety laboratory.</p><p><b>METHODS</b>first, two groups owning different experience simulated the course of accidental spills and splashes by recombinant adenovirus (rADV) which expressed green fluorescence protein (GFP), the GFP signal were observed in 96 well cell plate after spills appeared; Second, the routine two heights (75 cm and 110 cm) and capacity (1 ml, 1.5 ml, 4 ml and 8 ml) of virus were chose to simulate the experiment of unexpected dropping.</p><p><b>RESULTS</b>First, the positive quantity of the first group owning 5 years' experience is much less than the second group owning 2 years' work experience, the former was 7 positive wells, the latter was 81 positive when they used the pipette to operation. Second, when the unclosed test tubes (1 ml, 1.5 ml, 4 ml and 8 ml recombinant virus) inadvertently dropped, the largest spill distance was 0.92 m, 1.57 m, 2.63 m and2.68 m respectively.</p><p><b>CONCLUSION</b>The better experience is important to make sure safety when we make infectious material; the contaminated distance increased with the amount of recombinant virus material.</p>

Molecular interaction between PrP protein and the signal protein 14-3-3 beta / 病毒学报

The molecular interaction between PrP and 14-3-3 beta and the possible interactional domain between two proteins were studied by co-immunoprecipitation, pull down and FRET assays. The results showed that PrP protein could interact with 14-3-3 beta in vitro and in vivo. The domain which responded for the interaction was located at C-terminal of PrP (amino acid residues 106 to 126). This study of the interaction between PrP and 14-3-3 protein further provided the insight into the potential role of 14-3-3 in the biological function of PrP and the pathogenesis of prion disease.

Alternations of tau protein and its phosphorylated profiles in the experimental hamsters infected by scrapie agents 263K and 139A / 病毒学报

In human prion diseases, phosphorylated-tau deposition has been described in a rare genetic form, Gerstmann-Straussler-Scheinker disease, but is not considered part of the neuropathological picture of Creutzfeldt-Jakob disease. To investigate the possible changes of tau and phosphorylated tau (Ser396/Ser404) in transmissible spongiform encephalopathies (TSEs), the expressions and transcriptions of above biological factors in the brain tissues of 263K- and 139A-infected hamsters were evaluated by Western blots and Real Time PCR, respectively, followed by quantitative analyses of immunoblot images and relative transcriptional levels compared with normal animals. The contents of total tau increased, but phosphorylated tau at Ser396 and Ser404 decreased, regardless of the types of scrapie agents and clinical incubations. Transcriptions of two tau isoforms were also markedly increased. These findings suggested that dephosphorylation of tau at Ser396/Ser404 was a illness-correlative phenomenon in TSEs. Alterations of tau and phosphorylated tau (Ser396/Ser404) were either intermediate or consequent events in TSE pathogenesis and proposed the potential linkage of these bioactive proteins with the pathogenesis of prion diseases.

Establishment of a stable PrP(Sc) panel from brain tissues of experimental hamsters with scrapie strain 263K / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To establish a stable PrP(Sc) panel from brain tissues of experimental hamsters infected with scrapie agent 263K for evaluating diagnostic techniques of human and animals' prion diseases.</p><p><b>METHODS</b>Thirty brain tissue samples from hamsters intracerebrally infected with scrapie strain 263K and another 30 samples from normal hamsters were selected to prepare 10%, 1%, and 0.5% brain homogenates, which were aliquoted into stocks. PrP(Sc) in each brain homogenate was determined by proteinase K digestions followed by Western blot assay and partially by immunohistochemistry. Stability and glycoforms of PrP(Sc) were repeatedly detected by PrP(Sc)-specific Western blots in half a year and 3 years later.</p><p><b>RESULTS</b>PrP(Sc) signals were observed in all 10% brain homogenates of infected hamsters. Twenty out of 30 stocks and 19 out of 30 stocks were PrP(Sc) positive in 1% and 0.5% brain homogenatesof infected hamsters, respectively. Twenty-seven out of 30 stocks presented three positive bands in 10% brain homogenates, whereas none of 1% and 0.5% homogenates contained 3 bands. The detection of PrP(Sc)-specific signals stored in half a year and 3 years later demonstrated that the ratio of PrP(Sc) positive samples and glycoforms was almost unchanged. All normal hamsters' brain homogenates were PrP(Sc) negative.</p><p><b>CONCLUSION</b>A PrP(Sc) panel of prion disease can be established, which displays reliably stable PrP(Sc)-specific signals and glycoforms.</p>

Study on the characteristics of patients with Creutzfeldt-Jakob disease under 2008 surveillance data in China / 中华流行病学杂志

Objective To describe the epidemiological and clinical characteristics of Creutzfeldt-Jakob disease (CJD) in China. Methods Clinical and epidemical data on patients from China CJD surveillance network was analyzed. Blood and cerebral spinal fluid (CSF) specimens from these patients were collected. Western blot assay was used to detect 14-3-3 protein in CSF, PCR and sequencing assay were used for analyzing the polymorphism of 129 amino acid and mutation of PRNP gene. Results A total number of 31 probable and 11 possible sporadic CJD patients were identified. Additionally, one patient with Gerstmann-Straussler-Scheinker syndrome (GSS) and 2 familial CJD cases were identified. No geographic- or occupational-related events were observed among these cases. The mean age of onset on the probable or possible CJD patients were 56.7 and 57.4 years old, with sex ratios of the probable CJD patients as 8:9 and the possible one as 5:6 respectively. Rapid progressive dementia was the main foremost symptom, presenting in 33.3% of the CJD patients. Probable CJD patients showed more clinical manifestations than those possible ones. Conclusion Geography distribution, occupation, ratio of gender and the mean onset age of the CJD eases in 2008 were consistent with the characteristics of the sporadic CJD.

Investigating the risk of two model virus in the laboratory / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To study the survival time of recombination rival in environment and inactivation ability of different disinfectant and ultraviolet radiation against virus.</p><p><b>METHODS</b>NC membranes absorbed the recombinant adenovirus (rADV) or herpes simplex virus (rHSV) with green fluorescence protein (GFP) were laid, or immersed in various concentration of different disinfectants such as ethanol, sodium hypochlorite, lysol and geramine and then taked out them every 15 min, or exposed under ultraviolet radiation, then the NC membranes were adsorbed 1 h in cell, 37 degrees C 5% CO2 48 h. The results were observed under the fluorescence microscope.</p><p><b>RESULTS</b>(1) the average survival time of rHSV under environment is less than 60 min, rADV is almost up to 2 h. (2) The infection ability of rHSV and rADV was inactived 15 min by both ethanol (100%, 70% and 50%) and sodium hypochlorite (5%, 2.5% and 1.25%). (3) Two virus can be killed by 0.1% bromogeramine. (4) Both 5% and 2.5% lysol, but rADV can not lost the infection on Vero Cell until 75 min by 1.25% Lysol. (5) The rHSV was inactivated under ultraviolet radiation, but rADV was not.</p><p><b>CONCLUSION</b>The survival time of is different from both envelope rival and the no-envelope viral under nature environment and the inactivate ability of disinfectant also is different between two model virus; Disinfectant should be choose according to virus type.</p>

Establishment of a prion disease PrP(Sc) panel from the brain tissues of experimental hamsters infected with scrapie agent 263K / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To establish a prion disease PrP(Sc) panel from the brain tissues of experimental hamsters and to address the stability of the panel conserved under the specific condition, for evaluating the diagnostic techniques of human and animal's prion diseases.</p><p><b>METHODS</b>30 brain tissues of hamsters infected with scrapie strain 263K intracerebrally and 30 ones of normal hamsters were enrolled in this panel. Each brain sample was prepared to 10%, 1% and 0.5% homogenates and aliquoted into stocks. The presences of PrP(Sc) in each brain sample were evaluated with PrP-specific Western Blots and partially with immunohistochemistry, and the stability of PrP(Sc) signals in each sample were repeatedly assessed half a year later and 3 years later.</p><p><b>RESULTS</b>PrP(Sc) signals were detected in all stocks of 10% brain homogenates from infected hamsters, 26 out of 30 stocks of 1% homogenates and 19 out of 30 stocks of 0.5% homogenates. The assessments of PrP(Sc) signals in all samples half-year and three years later demonstrated almost unchanged. All homogenates of brain tissues of normal hamsters were PrP(Sc) negative.</p><p><b>CONCLUSION</b>A prion disease PrP(Sc) panel of the brain tissues, which includes 90 PrP(Sc) positive stocks and PrP(Sc) negative ones, was successfully established, with a reliable stability of PrP(Sc) signals.</p>

Analyses of the expressions of GFAP in the brain tissues of hamsters infected with various amounts of scrapie strain 263K at terminal stage / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To investigate whether gliosis in the brain tissues of the hamsters infected with various amounts of scrapie strain 263K is correlated with the inoculation doses or the incubation times.</p><p><b>METHODS</b>The total values of glial fibrillary acidic protein (GFAP) in brains were evaluated by Western Blots and the GFAP-stained cells were detected by immunohistochemistry (IHC). The characteristics of GFAP distributions among various groups were defined by quantitive and statistic analyses.</p><p><b>RESULTS</b>Compared with the brain tissues of normal hamsters, remarkably higher total GFAP levels and more GFAP-stained cells were observed in the brain tissues of infected ones, howbeit, no significant difference was addressed among the infected groups.</p><p><b>CONCLUSION</b>Inoculations of various amounts of scrapie strain 263K into experimental hamsters intracerebrally induced the similar patterns of gliosis in the brains at the clinically terminal stage, regardless of infectious doses and incubation times.</p>

The construction and identification of the PRNP vectors with ubiquitin or the lysosome-targeting signal / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To evaluate PrP expression characteristic of PRNP nucleic acid vaccine vector with ubiquitin or the lysosome-targeting signal.</p><p><b>METHODS</b>The gene of ubiquitin and lysosome-targeting signal were ligated to PRNP and pcDNA3.1 vector that is, pcDNA3.1-UPrP and pcDNA3.1-PrPL were constructed. The expression characteristics of PrP with two signals were evaluated by Western Blot and the localization was observed by indirect immune fluorescence.</p><p><b>RESULTS</b>The protein expressed by pcDNA3.1-UPrP and pcDNA3.1-PrPL with ubiquitin and lysosome-targeting signal can be recognized by prion-specific antibody. The protein has three glycosylation molecules form as native PrP.PrP with ubiquitin was degraded gradually with time extension,whereas quantity of PrP with lysosome signal reduced in 48 h after transfection. The protein with two location signals can direct fusion proteins to cytoplasm.</p><p><b>CONCLUSION</b>The PRNP vectors with ubiquitin or the lysosome-targeting signal were constructed and expressed in eukaryocyte successfully. There will be one of good foundation on PRNP nucleic acid vaccine.</p>