Phylo-geographic analysis of whitefly on the basis of mitochondrial cytochrome oxidase 1 gene / Análise filogeográfica da mosca-branca com base no gene citocromo oxidase 1 mitocondrial

Bemisia tabaci is a species complex that causes damage to its broad range of plant hosts through serious feeding. It transmits plant viruses of different groups to important agricultural crops. Some important cash crops of Pakistan are sugar cane, rice, tobacco and seed oil. It shows high genetic variability and is differentiated as races or biotypes. Biotypes are, biotype Q, biotype B, biotype B2, biotype M, biotype L, biotype A, biotype H, biotype C, biotype K, biotype N, biotype R, biotype E, biotype P, biotype J, biotype S, biotype AN. Although the current report based on the Bayesian study of mitochondrial cytohrome oxidase gene1 (CO1) DNA sequences has classified the different populations of whiteflies into twelve genetic groups which are Mediterranean, Sub-Saharan Africa silverleafing, Indian Ocean, Asia II, Asia I, Australia, New World, Italy, China, Sub-Saharan Africa non-silverleafing, Mediterranean/Asia Minor/Africa and Uganda sweet potato. Begomoviruses is largest group of viruses transmitted by B. tabaci and cause major diseases of crops such as tomato and chili leaf curl disease, cassava mosaic disease; yellow mosaic disease of legumes and cotton leaf curl disease. The main objective of current study is to inculpate knowledge regarding genetic diversity of whitefly in cotton fields across Pakistan via analysis of partial DNA sequence of mitochondrial gene Cytochrom Oxidase I (mtCO1).

Bemisia tabaci é um complexo de espécies que causa danos a uma ampla gama de hospedeiros vegetais por meio de alimentação séria. Ele transmite vírus de plantas de diferentes grupos para importantes safras agrícolas. Algumas safras comerciais importantes do Paquistão são cana-de-açúcar, arroz, tabaco e óleo de semente. Apresenta alta variabilidade genética e é diferenciado em raças ou biótipos. Os biótipos são: biótipo Q, biótipo B, biótipo B2, biótipo M, biótipo L, biótipo A, biótipo H, biótipo C, biótipo K, biótipo N, biótipo R, biótipo E, biótipo P, biótipo J, biótipo S, biótipo AN . Embora o relatório atual baseado no estudo bayesiano das sequências de DNA do gene 1 da oxidase do citocromo mitocondrial (CO1) tenha classificado as diferentes populações de moscas-brancas em doze grupos genéticos, que são Mediterrâneo, África Subsaariana com folha de prata, Oceano Índico, Ásia II, Ásia I, Austrália, Novo Mundo, Itália, China, África Subsaariana sem folha prateada, Batata-doce Mediterrâneo / Ásia Menor / África e Uganda. Os begomovírus são o maior grupo de vírus transmitidos por B. tabaci e causam as principais doenças de culturas, como a doença do cacho do tomate e da pimenta-malagueta, doença do mosaico da mandioca, doença do mosaico amarelo de leguminosas e doença do enrolamento da folha do algodão. O principal objetivo do presente estudo é inculpar conhecimento sobre a diversidade genética da mosca-branca em campos de algodão em todo o Paquistão por meio da análise da sequência parcial de DNA do gene mitocondrial Citocromo Oxidase I (mtCO1).

Phylo-geographic analysis of whitefly on the basis of mitochondrial cytochrome oxidase 1 gene

Abstract Bemisia tabaci is a species complex that causes damage to its broad range of plant hosts through serious feeding. It transmits plant viruses of different groups to important agricultural crops. Some important cash crops of Pakistan are sugar cane, rice, tobacco and seed oil. It shows high genetic variability and is differentiated as races or biotypes. Biotypes are, biotype Q, biotype B, biotype B2, biotype M, biotype L, biotype A, biotype H, biotype C, biotype K, biotype N, biotype R, biotype E, biotype P, biotype J, biotype S, biotype AN. Although the current report based on the Bayesian study of mitochondrial cytohrome oxidase gene1 (CO1) DNA sequences has classified the different populations of whiteflies into twelve genetic groups which are Mediterranean, Sub-Saharan Africa silverleafing, Indian Ocean, Asia II, Asia I, Australia, New World, Italy, China, Sub-Saharan Africa non-silverleafing, Mediterranean/Asia Minor/Africa and Uganda sweet potato. Begomoviruses is largest group of viruses transmitted by B. tabaci and cause major diseases of crops such as tomato and chili leaf curl disease, cassava mosaic disease; yellow mosaic disease of legumes and cotton leaf curl disease. The main objective of current study is to inculpate knowledge regarding genetic diversity of whitefly in cotton fields across Pakistan via analysis of partial DNA sequence of mitochondrial gene Cytochrom Oxidase I (mtCO1).

Resumo Bemisia tabaci é um complexo de espécies que causa danos a uma ampla gama de hospedeiros vegetais por meio de alimentação séria. Ele transmite vírus de plantas de diferentes grupos para importantes safras agrícolas. Algumas safras comerciais importantes do Paquistão são cana-de-açúcar, arroz, tabaco e óleo de semente. Apresenta alta variabilidade genética e é diferenciado em raças ou biótipos. Os biótipos são: biótipo Q, biótipo B, biótipo B2, biótipo M, biótipo L, biótipo A, biótipo H, biótipo C, biótipo K, biótipo N, biótipo R, biótipo E, biótipo P, biótipo J, biótipo S, biótipo AN . Embora o relatório atual baseado no estudo bayesiano das sequências de DNA do gene 1 da oxidase do citocromo mitocondrial (CO1) tenha classificado as diferentes populações de moscas-brancas em doze grupos genéticos, que são Mediterrâneo, África Subsaariana com folha de prata, Oceano Índico, Ásia II, Ásia I, Austrália, Novo Mundo, Itália, China, África Subsaariana sem folha prateada, Batata-doce Mediterrâneo / Ásia Menor / África e Uganda. Os begomovírus são o maior grupo de vírus transmitidos por B. tabaci e causam as principais doenças de culturas, como a doença do cacho do tomate e da pimenta-malagueta, doença do mosaico da mandioca, doença do mosaico amarelo de leguminosas e doença do enrolamento da folha do algodão. O principal objetivo do presente estudo é inculpar conhecimento sobre a diversidade genética da mosca-branca em campos de algodão em todo o Paquistão por meio da análise da sequência parcial de DNA do gene mitocondrial Citocromo Oxidase I (mtCO1).

Construction and evaluation of reference standards for detection and quantification of Klebsiella pneumoniae using real-time PCR / 西安交通大学学报·英文版

Objective: To construct reference standards for detection and quantification of Klebsiella pneumoniae (K. pneumoniae) with SYBR Green I-based real-time PCR assay. Methods: Primers were designed based on the published sequence of the phoE gene of K. pneumoniae. The standard was prepared by cell culture, PCR and T-A clone methods, and was identified by colony PCR and DMA sequencing. Results: The standard curve showed a very good linear negative regression between threshold cycle (Ct) and Log starting quantity of copy number. The detection range was from 5.2 to 5.2 × 106 copies per reaction, and the detection limit was 6 copies per reaction. The coefficients of variance (CVs) of three parallel experiments were in the range of 0.05 %-0.91 %. Conclusion: The reference standards have high stability and reproducibility. They can be used in the quantitative detection of K. pneumoniae.

Construction and evaluation of reference standards for detection and quantification of Klebsiella pneumoniae using real-time PCR / 药物分析学报

Objective To construct reference standards for detection and quantification of Klebsiella pneumoniae (K.pneumoniae) with SYBR Green I-based real-time PCR assay. Methods Primers were designed based on the published sequence of the phoE gene of K.pneumoniae. The standard was prepared by cell culture, PCR and T-A clone methods, and was identified by colony PCR and DNA sequencing. Results The standard curve showed a very good linear negative regression between threshold cycle (Ct) and Log starting quantity of copy number. The detection range was from 5.2 to 5.2×106 copies per reaction, and the detection limit was 6 copies per reaction. The coefficients of variance (CVs) of three parallel experiments were in the range of 0.05%-0.91%. Conclusion The reference standards have high stability and reproducibility. They can be used in the quantitative detection of K.pneumoniae.

An Efficient Method to Prepare PCR Cloning Vectors

An improved procedure for preparing PCR cloning vectors was developed. This procedure includes the incorporation of adapters to create XcmI restriction enzyme sites in pBluescript II SK(+) vectors, digestion with XcmI followed by further digestion of the small fragment produced by XcmI digestion with additional enzymes, and purification with PCR purification kits. Using this procedure, PCR cloning vectors with high ligation efficiencies and low blue or false-positive colonies were obtained.

A new DNA-cloning vector for Haemophilus influenzae Rd.

A new, high-efficiency, DNA-cloning vector pJ1-8 was derived in two steps from the chimeric plasmid pD7 consisting of RSF 0885 (ampr) and Haemophilus influenzae chromosomal DNA. pJl-8 has only one EcoRI site and a molecular weight of only 2.5 × 106. No detectable ampr transformation was obtained with pJl-8 DNA. However, ampr transformation increases markedly if Haemophilus influenzae chromosomal DNA segments are spliced into it, providing a very facile assay for detecting inserts.

Construction of cloning vector of c-myc-TFF2 fusion gene by PCR based gene assembly / 西安交通大学学报(医学版)

Objective To assemble fusion gene of c-myc tagged human trefoil factor family 2(hTFF2)(in vitro) and construct a cloning vector of this fusion gene.Methods Based on amino acid sequence of hTFF2 and codon of lactococcus lactis,cDNA of htff2 sequence was designed and extended at their 5' ends with a sequence encoding the c-myc tag;and the sequence of fusion gene c-myc-htff2 was designed.According to restriction enzyme sites of pBluescript Ⅱ sk(+),the SalⅠ and BamHⅠ were arranged at 5′ and 3′ ends of the fusion gene respectively.Instructed by DNAWORKS program,the fusion gene sequence of c-myc-htff2 was designed as 14 oligonucleotides that overlapped each other.The target gene fragment of cmyc-htff2 fusion gene was obtained by the means of polymerase chain reaction(PCR) based gene assembly.Then,the c-myc-htff2 was subcloned into vector of pBluescript II sk(+) for identification of the fusion gene by restricting enzyme excision and DNA sequencing.Results Assembly of c-myc-TFF2 fusion gene(in vitro) and construction of pBS-TFF2 had been completed successfully;and DNA sequencing showed that the sequence of synthetic gene accorded with the expectation.Conclusion With the help of DNAWORKS program,the design and assembly of oligonucleotides (in vitro) is effective to synthesize a target gene,and the cloning vector of c-myc-htff2 fusion gene has been successfully constructed.

THE CONSTRUCTION OF HYBRID pBR322: RSF1010 / 第二军医大学学报

The plasmids pBR322 and RSF1010, extracted from E.coli HB802 and E. coli C600 respectively were treated with restrictive endonuclease EcoRI and ligated with T4 DNA ligase. C600 strains were then transformed with the hybrid plasmid, and thus 253 transformants were obtained.The figures of gel elctrophoresis and electromicrograph proved that PSMM1 was a pBR322 : RSF1010 composite hybrid.In E. coli C600 PSMM1 obtained resistance against AP, Tc and Sm.The orientation of pBR322 and RSF1010 in hybrid PSMM1 and the failure of introduction of this hybrid into P.Putida AC10 were discussed