Stable expression of shRNA for the control of recombinant adenovirus replication

Preventing the replication of adenovirus could have practical uses, such as controlling infection with wild-type virus or in applications involving recombinant vectors. Mainly transient methods have been used to inhibit adenovirus replication, including siRNA or drugs. Here, we tested whether stable expression of shRNA designed to target hexon, Iva2, or pol can inhibit the replication of a recombinant adenoviral vector, Ad-LacZ (serotype 5, E1/E3 deleted), in 293T cells. Significant knockdown correlating with reduced Ad-LacZ replication was achieved only when hexon was targeted. Cell sorting and isolation of cellular clones further accentuated knockdown of the hexon transcript, reduced protein levels by more than 90%, and diminished adenovirus production. As visualized by transmission electron microscopy, the cellular clone expressing the hexon-specific shRNA yielded 89.2% fewer particles compared to the parental 293T cells. Full scale production followed by purification revealed a 90.2% reduction in Ad-LacZ biological titer. These results support the notion that stable expression of shRNA can be used as a means to control adenovirus replication.

Integrator Complex: Versatile Players in Transcriptional Regulation / 中国生物化学与分子生物学报

The discovery of integrator complex (INT) expanded our understanding of noncoding U-rich small nuclear RNA (U snRNA) maturation and transcriptional regulation, and has revived the research boom in related fields. This complex consists of at least 14 subunits, weighting over 1. 4 MD. On one hand, it functions by the cleavage of transcripts; on the other hand, it interacts with protein phosphatase 2A (PP2A) and dephosphorylates the C-terminal repeat sequence of RNA polymerase II (Pol II), thus regulating the transcriptional activity of RNA Pol II, and functions in the production of various RNAs (messenger RNA, small nuclear RNA, enhancer RNA, etc.). INT is recruited to the CTD during transcriptional initiation stage and moves along the U small nuclear RNA (snRNA) gene during transcription. Upon the recognition of 3' maturation sequence element, its cleavage activity is triggered and the matured transcripts are released. Besides, it is also involved in many other processes, including protein-coding gene transcription pause-release, transcriptional elongation, regulation of enhancer RNA transcription, and DNA and RNA metabolism etc. Meanwhile, the significance of INT complex or its components in tumorigenesis, pathogenesis, and ontogeny has being gradually highlighted. Nevertheless, the structural and compositional studies just took a new breakthrough recently. The 14 subunits that make up the complex are characterized by a large number of α-helices, which are further assembled into a huge transcription regulation machine based on the formation of functional modules. This article will mainly discuss about the composition, structural characteristics, functional studies, disease-association, and problem outlook of the integrator complex.

An update and perspectives on the use of promoters in plant genetic engineering

Genetically engineered plants have varied applications in agriculture for enhancing the values of food and feed.Genetic engineering aims to introduce selected genetic regions with desirable traits into target plants for bothspatial and temporal expressions. Promoters are the key elements responsible for regulating gene expressionsby modulating the transcription factors (TFs) through recognition of RNA polymerases. Based on theirrecognition and expression, RNA polymerases were categorized into RNA pol II and pol III promoters.Promoter activity and specificity are the two prime parameters in regulating the transgene expression. Since theuse of constitutive promoters like Cauliflower mosaic virus (CaMV) 35S may lead to adverse effects on nontarget organisms or ecosystem, inducible/tissue specific promoters and/or the RNA pol III promoters providemyriad opportunities for gene expressions with controlled regulation and with minimum adverse effects.Besides their role in transgene expression, their influence in synthetic biology and genome editing are alsodiscussed. This review provides an update on the importance, current prospects, and insight into the advantagesand disadvantages of promoters reported thus far would help to utilize them in the endeavour to developnutritionally and agronomically improved transgenic crops for commercialization.

Origin of RNA Polymerase II pause in eumetazoans: Insights from Hydra

Multicellular organisms have evolved sophisticated mechanisms for responding to various developmental,environmental and physical stimuli by regulating transcription. The correlation of distribution of RNAPolymerase II (RNA Pol II) with transcription is well established in higher metazoans, however genome-wideinformation about its distribution in early metazoans, such as Hydra, is virtually absent. To gain insights intoRNA Pol II-mediated transcription and chromatin organization in Hydra, we performed chromatin immunoprecipitation (ChIP)-coupled high-throughput sequencing (ChIP-seq) for RNA Pol II and Histone H3. Strikingly, we found that Hydra RNA Pol II is uniformly distributed across the entire gene body, as opposed to itscounterparts in bilaterians such as human and mouse. Furthermore, correlation with transcriptome datarevealed that the levels of RNA Pol II correlate with the magnitude of gene expression. Strikingly, thecharacteristic peak of RNA Pol II pause typically observed in bilaterians at the transcription start sites (TSSs)was not observed in Hydra. The RNA Pol II traversing ratio in Hydra was found to be intermediate to yeastand bilaterians. The search for factors involved in RNA Pol II pause revealed that RNA Pol II pausingmachinery was most likely acquired first in Cnidaria. However, only a small subset of genes exhibited thepromoter proximal RNP Pol II pause. Interestingly, the nucleosome occupancy is highest over the subset ofpaused genes as compared to total Hydra genes, which is another indication of paused RNA Pol II at thesegenes. Thus, this study provides evidence for the molecular basis of RNA Pol II pause early during theevolution of multicellular organisms.

ISSR analysis on genetic diversity of Ferula syreitschikowii in Xinjiang province / 中草药

Objective: To study the genetic relationship and genetic diversity of Ferula syreitschikowii germplasm resources in Xinjiang province. Methods: A total of 96 F. syreitschikowii germplasm resources of Xinjiang province were used as experimental materials. Fifteen primers selected from 33 ISSR primers could obtain high polymorphism and reproducibility bands for ISSR molecular marker analysis. Results: Total by 292 DNA bands were amplified including 289 polymarphic bands, counting for 99.01%. Genetic similarity analysis showed F. syreitschikowii germplasm resources of Xinjiang province with high genetic diversity. Through the statistical software analysis, the average number of effective alleles was 1.725 6, the average Nei's genetic diversity index was 0.4048, the average Shannon's information index was 0.587 9, and the genetic similarity coefficients among tested clones ranged from 0.500 0 to 0.828 7. These 96 germplasm resources were divided into two groups and 13 subgroups by UPGMA analysis, which could distinguish the germplasm resources from different sources. Conclusion: From the molecular level, it reveals the relationship between the geographical distribution of F. syreitschikowii and the germplasm resources and higher genetic diversity of germplasm resources, which provids basis for identification of F. syreitschikowii varieties.

Plant active LTR retrotransposons: a review / 生物工程学报

Long terminal repeat (LTR) retrotransposons are mobile DNA sequences that ubiquitously exist in eukaryotic genomes. They replicate themselves in the genome by copy-paste mechanism with RNA as medium. In higher plants, many active LTR retrotransposons have been applied to analyze molecular marker technology, genetic tagging, insertion mutation and gene function. Here, we systematically review the characteristics of plant active LTR retrotransposons, including their structures, copy numbers and distributions. We further analyzed the gag (group-specific antigen) and pol (polymerase) sequence features of different plants active LTR retrotransposons and the distribution patterns of the cis-acting elements in LTR regions. The results show that autonomous active LTR retrotransposons must contain LTR regions and code Gag, Pr, Int, Rt, Rh proteins. Both LTR regions are highly homologous with each other and contain many cis-regulatory elements; RVT and RNase_H1_RT domain are essential for Rt and Rh protein respectively. These results provide the basis for subsequent identification of plant active LTR retrotransposons and their functional analysis.

XRCC1 rs25487 locus genetic polymorphisms and susceptibility to nonobstructive azoospermia in Hui minority ethnic population of Shaanxi province / 西安交通大学学报(医学版)

Objective To explore the association between X-ray repair cross complementing group 1 (XRCC1)gene rs25487 locus polymorphisms and nonobstructive azoospermia in Hui minority ethic population of Shaanxi Province.Methods We used polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP)method for genotyping at XRCC1 gene rs25487 locus in 79 patients with nonobstructive azoospermia and 82 healthy male controls in Hui minority ethic population of Shaanxi Province.Then we analyzed the association be-tween XRCC1 gene rs25487 locus and nonobstructive azoospermia.Results Compared with GG genotype,GA, AA and GA + AA genotypes demonstrated a significantly increased risk for nonobstructive azoospermia (OR =2.286,95% CI 1.1 5 1-4.539;OR =2.202,95% CI 0.753-6.439;OR =2.271,95% CI 1.1 71-4.403),respec-tively.Meanwhile,the A allele frequency was significantly higher in azoospermic patients than in controls (OR =1.582,95% CI 1.005-2.492,P =0.047).Conclusion G→A in XRCC1 gene rs25487 locus is correlated with nonobstructive azoospermia in Hui minority ethic population of Shaanxi province.

Plant regulators and invertase activity in sugarcane at the beginning of the harvest season / Reguladores vegetais e atividade de invertases em cana de açúcar no início da safra

Invertases play an essential role in partitioning photosynthates between storage and growth. The objective of this study was to evaluate the activity of acid and neutral invertases and the role they play in controlling the accumulation of sucrose in sugarcane as a result of the application of plant regulators in the beginning of the cropping season.A randomized block experimental design was adopted, with five replicates.The treatments consisted in the application of three plant regulators of the class of growth inhibitors (Sulfomethuron-methyl - 20g ha-1, Glyphosate - 0.4L ha-1, and Compounds from organic carboxylic radicals + Glyphosate - 1L ha-1 + 0.15L ha-1), in addition to a control (natural ripening).The acid and neutral invertase levels are affected in different ways and intensities, due to the active principle used as ripening agent and to the weather conditions.In sugarcane variety RB85-5453, with the conditions described in this experiment, it is suggested high levels of soluble acid invertase in relation to levels of neutral invertase; however, the first was characterized by high sucrose content in the stalks.Inverse correlation could be established for sugarcane variety RB85-5453 between soluble acid invertase levels and effective sucrose accumulation in the stalks.

Invertases desempenham um papel essencial no particionamento de fotossintatos entre armazenamento e crescimento. O objetivo deste estudo foi avaliar a atividade das enzimas invertases ácida e alcalina (neutra) e o papel que desempenham no controle do acúmulo de sacarose em cana-de-açúcar, como resultado da aplicação de reguladores vegetais no início da época de colheita. O delineamento experimental utilizado foi em blocos ao acaso, com cinco repetições. Os tratamentos consistiram na aplicação de três reguladores vegetais da classe dos inibidores de crescimento (Sulfometuron-metil - 20g ha-1, Glifosato - 0,4L ha-1, e Compostos de radicais carboxílicos orgânicos + Glifosato - 1L ha-1 + 0,15L ha-1), além do controle (maturação natural). Os níveis das invertases ácida e neutra são afetados de maneiras e intensidades distintas, devido ao princípio ativo utilizado como agente de maturação e das condições meteorológicas. Para a variedade RB85-5453, nas condições descritas neste experimento, sugerem-se níveis elevados de SAI em relação à NI;no entanto, a primeira foi caracterizada por um elevado teor de sacarose nos colmos. Correlação inversa pode ser estabelecida entre a atividade da invertase ácida e o acúmulo de sacarose.

Detection of DNA polymerase λ activity during seed germination and enhancement after salinity stress and dehydration in the plumules of indica rice (Oryza sativa L.).

DNA polymerase λ (DNA pol λ) is the only reported X-family DNA polymerases in plants and has been shown to play a significant role in dry quiescent seeds, growth, development and nuclear DNA repair. cDNA for DNA pol λ has been reported in Arabidopsis and japonica rice cultivar and has been characterized from E. coli expressed protein, but very little is known about its activity at protein level in plants. The enzymatic activity of DNA pol λ was studied in dry, imbibed and during different germination stages of indica rice IR-8 (salt sensitive) by in-gel activity assay to determine its physiological role in important stages of growth and development. The upstream sequence was also analyzed using plantCARE database and was found to contain several cis-acting elements, including light responsive elements, dehydration responsive elements, Myb binding sites, etc. Hence, 4-day-old germinating seedlings of IR29, a salt-sensitive, but high yielding indica rice cultivar and Nonabokra, a salt-tolerant, but low yielding cultivar were treated with water (control) or 250 mM NaCl or 20% polyethyleneglycol-6000 for 4 and 8 h. The protein was analyzed by in vitro DNA pol λ activity assay, in-gel activity assay and Western blot analysis. DNA pol λ was not detected in dry seeds, but enhanced after imbibition and detectable from low level to high level during subsequent germination steps. Both salinity and dehydration stress led to the enhancement of the activity and protein level of DNA pol λ, as compared to control tissues. This is the first evidence of the salinity or dehydration stress induced enhancement of DNA pol λ activity in the plumules of rice (Oryza sativa L.) cultivars.

Avaliação da segurança alimentar e nutricional de famílias beneficiárias do Programa Bolsa Família / Food security status of beneficiary families of the brazilian 'Bolsa Família' Program

Objetivo: Avaliaram-se par�metros socioecon�micos e seguran�a alimentar e nutricional (SAN) de fam�lias benefici�rias do Programa Bolsa Fam�lia (PBF) residentes em um munic�pio do norte de Minas Gerais. M�todos: Foram investigadas 150 fam�lias e obtidos dados sobre as suas condi��es socioecon�micas (idade do titular, n�vel de escolaridade, situa��o no mercado de trabalho, renda gasta com alimenta��o e n�mero de filhos). Mediu-se o n�vel de seguran�a alimentar utilizando-se a Escala Brasileira de Medida da Inseguran�a Alimentar, com 15 quest�es que refletem a inseguran�a alimentar em diferentes n�veis de intensidade. Aplicou-se o teste de Fisher para avaliar a associa��o entre seguran�a alimentar e as vari�veis socioecon�micas. Resultados: A grande maioria das fam�lias era jovem, empregada e com at� tr�s filhos menores de 18 anos; 60% gastavam menos de 160 reais mensais com alimenta��o. Observou-se que 72,0% dos entrevistados sofriam de inseguran�a alimentar, sendo 50,0% em um grau leve, 14,7% moderada e 7,3% grave. Os dados de inseguran�a alimentar n�o se associaram �s vari�veis socioecon�micas. Conclus�o: As fam�lias contempladas pelo Programa Bolsa Fam�lia ainda vivem em condi��es de vulnerabilidade social e a seguran�a alimentar e nutricional ainda n�o � garantida para essa popula��o.

Objective: The present study evaluated the socioeconomic and food and nutrition security (FNS) parameters of families assisted under the ?Bolsa Fam�lia? Program in a municipality of northern Minas Gerais state, Brazil. Methods: Data on socioeconomic conditions (age and educational level of the head of family, labor market position, expenses with food, and number of children) were collected from a sample of 150 families. Food security status was measured using the Brazilian Food Insecurity Scale with 15 questions that indicate different levels of food insecurity. Association between food security and socioeconomic variables were assessed by the Fisher?s exact test. Results: Most families were young, with employed parents, and up to 3 underage children; 60% spent less than R$160.00/month on food. It was possible to observe that 72.0% of the families investigated suffered with food insecurity: 50.0% with mild insecurity, 14.7% with moderate insecurity, and 7.3% with severe insecurity. Data on food insecurity were not associated with the socioeconomic variables assessed. Conclusion: The families assisted by the ?Bolsa Fam�lia? Program still live under social vulnerability and do not have their food and nutritional security ensured.

Association of HSP70 gene polymorphisms with type 2 diabetes mellitus in Hui and Han people in Ningxia / 军事医学

Results There was no significant difference in the distribution of genotypes and allele frequencies of HSP 70-1 and HSP70-hom between case group and control group (P>0.05), but there was significant difference in the distribution of genotypes and allele frequencies of HSP70-2 between the two groups(P0.05)or in the distribution of genotypes of HSP 70-2 between males and females in case group, but there was significant difference in the distribution of allele frequencies of HSP 70-2 in case group .There was no significant difference in the distribution of genotypes and allele frequencies of HSP 70-1 and HSP70-hom between males and females in case or control group .Logistic analysis confirmed that the WC, TG, TC, LDL-C, SBP, family history of diabetes and G allele were risk factors of T 2DM. Conclusion GG genotype and G allele of HSP 70-2 (+1267 ) SNP may be genetic markers for the susceptibility of type 2 diabetes.There is no significant difference in the distribution of genotypes and allele frequencies of HSP 70-1,HSP70-2 and HSP70-hom between Hui and Han nationalities .Family history of diabetes, LDL-C, TC, and G allele of HSP70-2 (+1267) SNP are the main risk factors of T2DM while HDL-C is a protective factor.

Staphylococcal chromosome mec genotyping and molecular epidemiology of hospi-tal-acquired methicillin-resistant Staphylococcus aureus / 中国感染与化疗杂志

Objective To investigate the staphylococcal chromosomal cassette mec (SCCmec)genotypes and molecular epidemi-ology of hospital-acquired MRSA.Methods A total of 26 non-duplicate MRSA isolates with the same resistant pattern were studied.SCCmec genotyping was analyzed by multiplex PCR.Repetitive element polymerase chain reaction (Rep-PCR)tech-nique was used to analyze the homology between these strains based on the DiversiLab system.Results The most common geno-type of these MRSA strains was SCCmec-III (84.6%).Two strains belonged to SCCmec-II and 1 SCCmec-IV.SCCmec-I strain was not identified.Based on the results of DiversiLab analysis,these MRSA strains were classified into 10 groups.The genetic similarity ranged from 40% to 100% among these SCCmec types.The two strains of SCCmec-II belonged to the same subtype.The similarity coefficient was higher than 90% for one strain of SCCmec-III subtype 1.The 4 strains of SCCmec-III subtype 3 were grouped into the same set with a similarity coefficient of > 95%.The MRSA strains of SCCmec-III subtype 2 was divided into 5 groups (similarity co-efficient > 90%).Conclusions SCCmec-III is the major genotype of MRSA isolates in our hospital.MRSA strains may spread in some wards.Clinicians and infection control department should pay close attention to this issue.

Processos de exclusão social e iniquidades em saúde: um estudo de caso a partir do Programa Bolsa Família, Brasil / Social exclusion and health inequity: a case study based on a cash distribution program (Bolsa Família) in Brazil

OBJETIVO: Compreender as repercussões do Programa Bolsa Família (PBF) e analisar seus efeitos nos processos de inclusão e exclusão social vividos pelas famílias pobres no Brasil, em especial sua potencialidade para enfrentar iniquidades em saúde. MÉTODOS: A investigação de abordagem qualitativa empregou a metodologia de estudo de caso com utilização das técnicas de observação participante, pesquisa documental e entrevistas semiestruturadas com famílias beneficiárias e ex-beneficiárias do PBF, além de gestores municipais. O estudo foi conduzido em um município de pequeno porte do estado do Rio de Janeiro, com elevado índice de exclusão social e cobertura de 100% da Estratégia Saúde da Família (ESF).A abordagem dosprocessos deinclusão e exclusão socialem suas dimensões econômica, social, política e cultural foi utilizada para orientar a coleta e análise dos dados. RESULTADOS: O programa favoreceu a inclusão social das famílias pobres, especialmente nas dimensões econômica e social, apesar de não promover as mudanças reivindicadas pelos beneficiários na esfera do trabalho. Os efeitos na dimensão política foram limitados pelo funcionamento inadequado das instâncias de participação social. Os entrevistados destacaram os efeitos positivos da ESF relacionados ao usufruto do direito à saúde, em particular a ampliação do acesso e utilização de serviços de saúde de atenção primária. No entanto, esses efeitos mostraram-se desvinculados do PBF. CONCLUSÕES: O trabalho aponta efeitos, limites e desafios do PBF para modificar os determinantes sociais produtores de iniquidades da saúde, a fim de que se alterem, de modo mais permanente, as dinâmicas de exclusão/inclusão social de famílias vivendo em situação de pobreza.

OBJECTIVE: To understand the impact of Bolsa Família (PBF), a federal cash transfer program, and to analyze its effects on social inclusion and exclusion processes experienced by low-income families in Brazil, with a focus on the program's potential to help overcome health inequity. METHODS: This qualitative investigation used a case study methodology including observant participation, review of documents, and semi-structured interviews with current and former PBF beneficiaries, as well as with the program's local managers. The study was conducted in a small city in the state of Rio de Janeiro with a high social exclusion index and 100% coverage by the Family Health Strategy (Estratégia Saúde da Família, ESF) program. The economic, political, social, and cultural dimensions of social exclusion and inclusion processes were used to guide data collection and analysis. RESULTS: The program facilitated social inclusion of low-income families, especially in the economic and social dimensions. Nevertheless, it did not produce the changes desired by the beneficiaries in the work dimension. The effects on the political dimension were limited by the insufficient social engagement of the PBF. The interviewees underscored the positive effects of the ESF, which allowed them to exercise their right to health by granting them wider access to primary health care services. However, these effects appeared to be unrelated to the PBF. CONCLUSIONS: The results reveal effects, limitations, and challenges of the PBF towards modifying the social determinants of health inequity, in order to promote more effective changes in the social exclusion/inclusion dynamics affecting low-income families.

Expression of rna polymerase IV and V in oryza sativa

RNA polymerase IV and V are principal players in the RdDM pathway, where their current study has shown interaction of several factors that control DNA silencing of intergenic regions and siRNA production. DNA silencing is an important process during cell differentiation, nuclear structure and viral control. However, RNA pol IV and V are yet to be study in model monocot systems like Oryza sativa that can provide further information on genetic silence mechanism in plats. We show the expression pattern of these polymerases in tissues extracts of Oryza sativa. Detectable amounts of these polymerases are found in specific adult plant tissues and particularly expressed during somatic embryogenesis but not during early stages of normal embryo development. The use of synthetic auxin leads to an induction of both RNA pol IV and V in scutellum tissue where nuclear localization may be required for genome reorganization and gene silencing.

Investigation of pol gene variation of HIV-1 epidemic strains after treatment with HARRT at Dehong prefecture and Kunming in Yunnan province / 中华检验医学杂志

Objective To investigate the variations in the pol region of HIV-1 strain in treatment failed patients in Yunnan province's Dehong prefecture and Kunming. Methods Blood samples were collected from 139 patients who experienced treatment failure ( HAART treatment ＞ 1 years and HIV-1 RNA Viral load ＞ 1 000 copies/ml). HIV-1 RNA was extracted from plasma, and nested-PCR was performed for amplification of PR and RT genes on the HIV-1 pol region. The PCR products were then sequenced and submitted to Stanford HIV Drug Resistance Database for comparison. The evolution tree was built up with MEGA 4. 1 system, combined with patients' demographics. Results The most prevalent mutation in Kunming patients were T215F/N/Y/I, M41L/M, and T69G/N/I/S/A/D, the mutation rates were 39%(24/62), 27% (17/62) and 27% (17/62) , respectively, which were higher than the corresponding mutations in the Dehong prefecture [16% ( 11/69), 13% (9/69) and 9% (6/69)]. The rate differences were statistically significant ( x2 = 8.646, 4.242 and 7. 909, all P ＜ 0.05 ). The most common HIV-1 pol region subtype in the Dehong patients were CRF01_AE subtype (32%, 22/69), followed by C subtype (25% ,17/69), and B subtype ( 19%, 13/69). Major subtypes in Kunming patients were 08_BC (60%,37/62 ), CRF01_AE subtype(21% , 13/62 ) and 07_BC ( 15% ,9/62). Conclusions Partial differences of the point mutations of the HIV-1 strain pol region and frequency of their occurrences exist among Dehong and Kunming patients, HIV-1 strains in Dehong prefecture for the NNRTIs mutations at the T215 Y/N/T, M41L and T69G/N/I/S/A/D are significantly higher than those in Kunming. Six isoforms are found respectively:CRF01_AE, B, C, BC, 08_BC and 07_BC from the epidemic strains of HIV-1 pol region subtype in Dehong and Kunming areas.

A second generation of RT-PCR assay for detection of human immunodeficiency virus type 1 (HIV-1) infection.

Aim A spesific and rapid diagnosis such as RT-PCR asay is the most needed to minimize transmission of HIV-1 infection. Therefore, in this study we developed the RT-PCR assay that was spesifi c against the gag gene of HIV-1. Methods The developed RT-PCR assay was evaluated against 46 specimens that were obtained from voluntary counseling and testing for HIV (VCT) in Rumah Sakit Umum Pemerintah (RSUP) Sanglah, Bali. To get the sensitivity and specifi city of RT-PCR assay, the results of assays were compared with the results of commercially serologic tests that were commonly used in Indonesia. Results The RT-PCR assay could detect 21 of 26 serologic test-positive specimens and showed 19 negative results of 20 serologic test-negative specimens. There was one specimen that was positive in RT-PCR but negative in serologic assay, which might depict a true yield at particular condition when the serologic assay was unable to detect. Five serologic positive-test specimens were negative by RT-PCR that was possibly caused by low detection level of the assay. Conclusion The RT-PCR assay is potential to be used for the detection of HIV-1 infection with a sensitivity and specificity of 80.8% and 95.0% respectively.

Pol-specific T lymphocyte responses in Chinese HIV-1 recombinant subtype B/C infectors at different stages of diseases / 中华微生物学和免疫学杂志

Objective To analyze the character of Pol-specific T lymphocyte responses and identify immunodominant region recognized in Chinese HIV-1 recombinant subtype B/C infectors at different stages of diseases. Methods Eleven Chinese HIV-1 recombinant subtype B/C infectors infected in 18 months, 25 which infected time more than 3 years and 10 HIV-1-seronegative healthy individuals were enrolled. HIV-1-specific T lymphocyte responses were analyzed by an IFN-γ ELISPOT assay against 249 overlapping peptides spanning HIV-1 Pol protein in the present study. Results Pol-specific T lymphocyte responses of IFN-γsecretion were identified in 8 (72.73%) out of 11 infectors infected in 18 months, the specific T lymphocytes are mainly targe-ted at six peptides which amino acid position from Pol 481 to 631 in reverse transcriptase region: Pol5581, Pol5582, Pol5587, Pol5609, Pol5610 and Pol5615. There was a negative correlation between the breadth of re-sponse and peripheral CD4+ T cell count (P=0.0212, r=-0.762) ; Responses were identified in 15 (60%) out of 25 chronic infectors, the specific T lymphocytes are mainly targeted at four peptides which amino acid po-sition from Pol 241 to 295: Pol5521, Pol5525, Pol5526, Pol5531 and another peptide: Pol5638 which amino acid position from Pol 708 to 722 in reverse transcriptase region. There was a positive correlation between the magnitude of Pol-specific IFN-γ secretion T lymphocyte responses and plasma viremia (P = 0.006 95 , r = 0.660) . None of the seronegative healthy individuals gave the positive responses. Conclusion Chinese HIV-1 recombinant subtype B/C infectors at different stages of diseases mainly recognized different re-gions of Pol.

Aislamiento del virus linfotrópico de células T humano tipo I de un paciente con paraparesia espástica tropical: primer reporte en Cuba / Isolation of human T cell lymphotropic virus type 1 from a patient affected by tropical spastic paraparesis: first report in Cuba

Introducción: la paraparesia espástica tropical o mielopatía asociada a HTLV-I (PET/MAH) es una afección neurológica crónica etiológicamente ligada al virus linfotrópico de células T humano tipo I (HTLV-I). Objetivo: realizar la confirmación serológica de la infección y el aislamiento del virus linfotrópico de células T humano tipo I, a partir de las células mononucleares de sangre periférica, de una paciente con un cuadro clínico compatible con paraparesia espástica tropical. Métodos: las células mononucleares de sangre periférica se cocultivaron durante 35 d. La detección viral se realizó por inmunofluorescencia específica de membrana y reacción en cadena de la polimerasa para los genes tax, pol y gag. Resultados: se confirmó la presencia de anticuerpos contra el virus linfotrópico de células T humano tipo I en el suero. Se observó un incremento del número de células positivas por inmunofluorescencia indirecta en el monitoreo secuencial y se detectó la presencia de ADN proviral en las células mononucleares de sangre periférica. Conclusiones: los resultados evidenciaron el aislamiento del virus y corroboraron, por primera vez en Cuba, la asociación entre el virus linfotrópico de células T humano tipo I y paraparesia espástica tropical.

Background: Tropical spastic paraparesis or HTLV-I-associated myelopathy (TSP/HAM) is a chronic neurological disease etiologically linked to human T-cell lymphotropic virus type I (HTLV-I). Objective: to serologically confirm infection by and isolation of HTLV-I from the peripheral blood mononuclear cells from a patient who presented with a clinical picture similar to that of tropical spastic paraparesis. Methods: peripheral blood mononuclear cells were co-cultured for 35 days. Viral detection by membrane- specific immunofluorescence and polymerase chain reaction of genes tax, pol and gag were performed. Results: The presence of antibody to human T cell lymphotropic virus type I in serum was confirmed. Indirect immunofluorescence in the sequential monitoring made it possible to observe an increase of the number of positive cells whereas proviral DNA was detected in the peripheral blood mononuclear cells. Conclusions: these results support the evidence of viral isolation and confirmed, for first time in Cuba, the association between HTLV-I and TSP.

Mapping the molecular characteristics of Brazilian human T-cell lymphotropic virus type 1 Env (gp46) and Pol amino acid sequences for vaccine design

This study was carried out to evaluate the molecular pattern of all available Brazilian human T-cell lymphotropic virus type 1 Env (n = 15) and Pol (n = 43) nucleotide sequences via epitope prediction, physico-chemical analysis, and protein potential sites identification, giving support to the Brazilian AIDS vaccine program. In 12 previously described peptides of the Env sequences we found 12 epitopes, while in 4 peptides of the Pol sequences we found 4 epitopes. The total variation on the amino acid composition was 9 and 17 percent for human leukocyte antigen (HLA) class I and class II Env epitopes, respectively. After analyzing the Pol sequences, results revealed a total amino acid variation of 0.75 percent for HLA-I and HLA-II epitopes. In 5 of the 12 Env epitopes the physico-chemical analysis demonstrated that the mutations magnified the antigenicity profile. The potential protein domain analysis of Env sequences showed the loss of a CK-2 phosphorylation site caused by D197N mutation in one epitope, and a N-glycosylation site caused by S246Y and V247I mutations in another epitope. Besides, the analysis of selection pressure have found 8 positive selected sites (w = 9.59) using the codon-based substitution models and maximum-likelihood methods. These studies underscore the importance of this Env region for the virus fitness, for the host immune response and, therefore, for the development of vaccine candidates.

Analysis of Plant TATA and TATA-less Promoters by Using Sequence and Structure Features / 生物化学与生物物理进展

Analysis of regular elements in promoter region is the base for elucidating the mechanism of gene transcription initiation.The TATA and the TATA-less promoters of plant RNA polymerase Ⅱ gene are chosen from the PlanPromDB.The GC bias,position structure conservation,nucleotide content and conservative motifs of sequences,position distribution of TATA box and conservation of correlation position are analyzed.Many specific regulars for the two types of promoters are found.These features can offer some help for revealing the transcription regulation of plant gene.A new prediction algorithm based on position-correlation weight matrix(PCWM) is proposed.The better discrimination results for two sort plant promoters are obtained by using score function.It is confirmed that the performance of position-correlation weight matrix(PCWM) is superior to single-base position weight matrix(PWM).