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Promega wizard pcr preps dna purification system
Wizard Pcr Preps Dna Purification System, supplied by Promega, used in various techniques. Bioz Stars score: 99/100, based on 204 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 99 stars, based on 204 article reviews
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wizard pcr preps dna purification system - by Bioz Stars, 2020-04
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Clone Assay:

Article Title: Mapping of Human Autoantibody Binding Sites on the Calcium-Sensing Receptor
Article Snippet: For surface expression, DNA fragments are required to be cloned in frame with the PelB leader peptide and the gene III phage coat protein present in pComb3 at the N- and C-terminal, respectively. .. The DNA fragments were purified using a Wizard PCR Preps DNA Purification System and treated with T4 DNA polymerase (Promega) according to the manufacturer's protocol to create blunt ends.

Article Title: A New Huanglongbing Species, "Candidatus Liberibacter psyllaurous," Found To Infect Tomato and Potato, Is Vectored by the Psyllid Bactericera cockerelli (Sulc) ▿
Article Snippet: This PCR product was then cloned using the GeneJET PCR cloning kit (Fermentas International Inc., Burlington, Ontario, Canada). .. Amplified DNA was cleaned using the Wizard PCR Preps DNA purification system (Promega, Madison, WI) and directly sequenced in both directions.

Article Title: Bovine torovirus: sequencing of the structural genes and expression of the nucleocapsid protein of Breda virus.
Article Snippet: .. Cloning and expression The PCR product containing the BRV N gene was purified using the Wizard PCR Preps DNA Purification System (Promega). .. Amplicons were then ligated to a pET28a(+) cloning vector (Novagen, Madison, WI) , and transformed into INVhF% One Shot cells (Invitrogen, Carlsbad, CA) as per the manufacturer's recommendations.

Amplification:

Article Title: A Deletion at the Mouse Xist Gene Exposes Trans-effects That Alter the Heterochromatin of the Inactive X Chromosome and the Replication Time and DNA Stability of Both X Chromosomes
Article Snippet: Each of these PCR products was amplified separately using Taq Polymerase (Promega, Madison, WI) from a region extending up to 14.7 kb upstream of the 5′ end of exon 2 of the Hprt gene and that is deleted in the Hprt -Δ allele used in this study ( ). .. The PCR products were gel extracted using the Wizard PCR Preps DNA purification system (Promega).

Article Title: Differential effects of a common splice site polymorphism on the generation of OAS1 variants in human bronchial epithelial cells.
Article Snippet: .. Isoform-specific RT-PCR products containing the target sequences, amplified with primers in Supplementary Table 1, were purified using the Wizard PCR Preps DNA Purification System (Promega, Fitchburg, WI, USA). .. Their copy numbers were calculated from the concentration of DNA determined by measuring absorbance at 260 nm.

Article Title: Cloning and expression of 3-deoxy-d-manno-oct-2-ulosonic acid α-ketoside hydrolase from oyster hepatopancreas
Article Snippet: .. The following materials were purchased from commercial sources: marker proteins for molecular weight and p I , Sephacryl S-300 HR, Amersham Pharmacia Biotech; Tris, bis-Tris, MU-α-Neu5Ac and other MU-glycosides, Sigma; phenylmethylsulfonyl fluoride, Pierce; Silica Gel-60 thin-layer chromatography (TLC) plates, EMD Chemicals, Darmstadt, Germany; Centricon-10 and Microcon (10 kDa M r cut-off) micro-concentrators, Amicon; E. coli BL21(DE3), E. coli DH5α, S. cerevisiae INVSc1, P. pastoris , CHO-S cell, pUC18, pET, pYES2, pcDNA3.1/ myc -His(A), pPICZ, 5′-Race System for Rapid Amplification of cDNA Ends (Version 2) and LipofectAMINE 2000, Invitrogen; E. coli Origami, Novagen; GeneAmp PCR System 9600, Perkin-Elmer; Wizard PCR Preps DNA Purification System and pGEM-T Easy Vector System, Promega; RNAlater, Ambion; YPD and SD yeast culture media, BD Bioscience; FastTrack 2.0 Kit, BD Smart Race cDNA Amplification Kit and BD Advantage 2 PCR Kit, Clontech; Thermoscript RT-PCR System, Gibco; Oligotex Direct mRNA midi/maxi Kit and Ni-NTA Superflow column, Qiagen; chemically defined natural Re-LPS from S. minnesota and E. coli D31m4, Rd-LPS from S. minnesota R7 LIST Biological Laboratories, Inc; LPS from Neisseria meningitides strains A1 and M992 were gifts of Dr. Chao-Ming Tsai, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland. .. The α-Kdo-cleaving activity was measured by fluorometric analysis according to the procedure of ) as described previously ( ).

Article Title: Molecular Characterization of Diphtheria Toxin Repressor (dtxR) Genes Present in Nontoxigenic Corynebacterium diphtheriae Strains Isolated in the United Kingdom
Article Snippet: The amplicons were purified using the Wizard PCR Preps DNA purification system (Promega Corp., Madison, Wis.) as specified by the manufacturer. .. Sequencing was performed on each strand with the primers used in the initial PCR amplification (DtxR 1F and DtxR 4R) and primers DtxR 2F (5′-CTGAGCGTCTGGAACAATCT-3′) and DtxR 3R (5′-ATCCAACACGGATGTCAGCAT-3′).

Article Title: A New Huanglongbing Species, "Candidatus Liberibacter psyllaurous," Found To Infect Tomato and Potato, Is Vectored by the Psyllid Bactericera cockerelli (Sulc) ▿
Article Snippet: .. Amplified DNA was cleaned using the Wizard PCR Preps DNA purification system (Promega, Madison, WI) and directly sequenced in both directions. .. With the 10F and 480R primers on tomato and potato plants, chloroplast DNA was preferentially amplified.

Article Title: Age-Dependent Demise of GNAS-Mutated Skeletal Stem Cells and "Normalization" of Fibrous Dysplasia of Bone
Article Snippet: This allowed for the selective amplification and sequencing of the mutant allele and is estimated to detect one mutant cell out of 200 cells. ( ) The target DNA (200–500 ng) was amplified in a standard 100-µl PCR reaction mixture using 2.5 U of Ampli Taq Gold polymerase (Perkin Elmer, Boston, MA, USA) and 1 µg of each primer, forward primer, 5′-GTTTCAGGACCTGCTT CGC -3′; reverse primer, 5′-GCAAAGCCAAGAGCGTGAG-3′, and 2 µg of the PNA sequence amino-terminal 5′- CGC TGCCGTGTC carboxy-terminal- 3′ (bold indicates overlap of the forward primer with the PNA sequence). .. The 325-bp PCR products were purified using the Promega Wizard PCR Preps DNA Purification System (Madison, WI, USA) and sequenced using a Perkin Elmer Applied Biosystem 377 Automated Sequencer, and the sequence was read from the 3′-5′ (reverse) strand.

Article Title: Prevalence and distribution of dermatophytosis lesions on cattle in Plateau State, Nigeria
Article Snippet: .. The amplified DNA was purified using Wizard PCR Preps DNA Purification System (Promega) and sequenced with the primers ITS-1 (5’-TCCGTAGGTGAACCTGCGG-3’) and ITS-4 (5’-TCCTCCGCTTATTGATATGC-3’) using the dye terminator method (Applied Biosystems 377 automatic sequencer). ..

Synthesized:

Article Title: Cloning and expression of 3-deoxy-d-manno-oct-2-ulosonic acid α-ketoside hydrolase from oyster hepatopancreas
Article Snippet: The enzyme substrates, MU-α-Kdo, MU-β-Kdo and MU-α-Kdn, were previously synthesized in our laboratory ( ; ). .. The following materials were purchased from commercial sources: marker proteins for molecular weight and p I , Sephacryl S-300 HR, Amersham Pharmacia Biotech; Tris, bis-Tris, MU-α-Neu5Ac and other MU-glycosides, Sigma; phenylmethylsulfonyl fluoride, Pierce; Silica Gel-60 thin-layer chromatography (TLC) plates, EMD Chemicals, Darmstadt, Germany; Centricon-10 and Microcon (10 kDa M r cut-off) micro-concentrators, Amicon; E. coli BL21(DE3), E. coli DH5α, S. cerevisiae INVSc1, P. pastoris , CHO-S cell, pUC18, pET, pYES2, pcDNA3.1/ myc -His(A), pPICZ, 5′-Race System for Rapid Amplification of cDNA Ends (Version 2) and LipofectAMINE 2000, Invitrogen; E. coli Origami, Novagen; GeneAmp PCR System 9600, Perkin-Elmer; Wizard PCR Preps DNA Purification System and pGEM-T Easy Vector System, Promega; RNAlater, Ambion; YPD and SD yeast culture media, BD Bioscience; FastTrack 2.0 Kit, BD Smart Race cDNA Amplification Kit and BD Advantage 2 PCR Kit, Clontech; Thermoscript RT-PCR System, Gibco; Oligotex Direct mRNA midi/maxi Kit and Ni-NTA Superflow column, Qiagen; chemically defined natural Re-LPS from S. minnesota and E. coli D31m4, Rd-LPS from S. minnesota R7 LIST Biological Laboratories, Inc; LPS from Neisseria meningitides strains A1 and M992 were gifts of Dr. Chao-Ming Tsai, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland.

TA Cloning:

Article Title: AgtA, the Dicarboxylic Amino Acid Transporter of Aspergillus nidulans, Is Concertedly Down-Regulated by Exquisite Sensitivity to Nitrogen Metabolite Repression and Ammonium-Elicited Endocytosis ▿, Is Concertedly Down-Regulated by Exquisite Sensitivity to Nitrogen Metabolite Repression and Ammonium-Elicited Endocytosis ▿ †
Article Snippet: TA cloning was carried out with the pGEM-T Easy vector system (Promega). .. DNA bands were purified from agarose gels with the Wizard PCR Preps DNA purification system (Promega).

Construct:

Article Title: Mapping of Human Autoantibody Binding Sites on the Calcium-Sensing Receptor
Article Snippet: To construct a CaSR cDNA fragment library in pComb3, full-length CaSR cDNA was prepared from pcCaSR-FLAG( ) by restriction of the plasmid with Kpn I (Promega) and Xba I (Promega). .. The DNA fragments were purified using a Wizard PCR Preps DNA Purification System and treated with T4 DNA polymerase (Promega) according to the manufacturer's protocol to create blunt ends.

Electrophoresis:

Article Title: Prevalence and distribution of dermatophytosis lesions on cattle in Plateau State, Nigeria
Article Snippet: Amplification products were separated by electrophoresis in 1.5% agarose gels incorporated with ethidium bromide and documented using gel documentation system (BIORAD Laboratories). .. The amplified DNA was purified using Wizard PCR Preps DNA Purification System (Promega) and sequenced with the primers ITS-1 (5’-TCCGTAGGTGAACCTGCGG-3’) and ITS-4 (5’-TCCTCCGCTTATTGATATGC-3’) using the dye terminator method (Applied Biosystems 377 automatic sequencer).

Incubation:

Article Title: Mapping of Human Autoantibody Binding Sites on the Calcium-Sensing Receptor
Article Snippet: The reaction was incubated at room temperature for 15 minutes and then terminated by the addition of 50 mM EDTA (pH 8.0). .. The DNA fragments were purified using a Wizard PCR Preps DNA Purification System and treated with T4 DNA polymerase (Promega) according to the manufacturer's protocol to create blunt ends.

Expressing:

Article Title: Mapping of Human Autoantibody Binding Sites on the Calcium-Sensing Receptor
Article Snippet: For surface expression, DNA fragments are required to be cloned in frame with the PelB leader peptide and the gene III phage coat protein present in pComb3 at the N- and C-terminal, respectively. .. The DNA fragments were purified using a Wizard PCR Preps DNA Purification System and treated with T4 DNA polymerase (Promega) according to the manufacturer's protocol to create blunt ends.

Article Title: Differential effects of a common splice site polymorphism on the generation of OAS1 variants in human bronchial epithelial cells.
Article Snippet: Isoform-specific RT-PCR products containing the target sequences, amplified with primers in Supplementary Table 1, were purified using the Wizard PCR Preps DNA Purification System (Promega, Fitchburg, WI, USA). .. We used the b-actin gene (primers in Supplementary Table 1) to normalize OAS1 expression.

Article Title: Bovine torovirus: sequencing of the structural genes and expression of the nucleocapsid protein of Breda virus.
Article Snippet: .. Cloning and expression The PCR product containing the BRV N gene was purified using the Wizard PCR Preps DNA Purification System (Promega). .. Amplicons were then ligated to a pET28a(+) cloning vector (Novagen, Madison, WI) , and transformed into INVhF% One Shot cells (Invitrogen, Carlsbad, CA) as per the manufacturer's recommendations.

CTG Assay:

Article Title: A Deletion at the Mouse Xist Gene Exposes Trans-effects That Alter the Heterochromatin of the Inactive X Chromosome and the Replication Time and DNA Stability of Both X Chromosomes
Article Snippet: The sequences of primers are: GCA AGC ATA AGG ACC AGA GC (412R), TTC CAC AAG AAA TAT TAC ACA AAA CA (412L), CCT AAC CAT TGA GCC GTC TT (605R), GGT CTC TGA ACT ACC AAT TGC AC (605L), GCA ATG ACA AAT GTT TTG TGG (609R), TGC TTA TTA GCA CAA GAC CTC AAG (609L), ATC ACC CTA TTC CCA GTG GA (850R), GCA GAT GAT AAG CTA TCC TTG AGA (850L), CAT CAC TGA GTC TTG CTG GTT T (1011R), CAA TTT AGG GGA AGG AAG CA (1011L), TGG TAG CTG GGC ATA AAA GC (1560R), AAT GGG AGA AAA GGC AGG AT (1560L), CAG GAA AGG GTG TGT GTG TG (1755R), and TAC GCT CTG GCA GTT TTC AA (1755L). .. The PCR products were gel extracted using the Wizard PCR Preps DNA purification system (Promega).

Transformation Assay:

Article Title: Bovine torovirus: sequencing of the structural genes and expression of the nucleocapsid protein of Breda virus.
Article Snippet: Cloning and expression The PCR product containing the BRV N gene was purified using the Wizard PCR Preps DNA Purification System (Promega). .. Amplicons were then ligated to a pET28a(+) cloning vector (Novagen, Madison, WI) , and transformed into INVhF% One Shot cells (Invitrogen, Carlsbad, CA) as per the manufacturer's recommendations.

Chloramphenicol Acetyltransferase Assay:

Article Title: A Deletion at the Mouse Xist Gene Exposes Trans-effects That Alter the Heterochromatin of the Inactive X Chromosome and the Replication Time and DNA Stability of Both X Chromosomes
Article Snippet: The sequences of primers are: GCA AGC ATA AGG ACC AGA GC (412R), TTC CAC AAG AAA TAT TAC ACA AAA CA (412L), CCT AAC CAT TGA GCC GTC TT (605R), GGT CTC TGA ACT ACC AAT TGC AC (605L), GCA ATG ACA AAT GTT TTG TGG (609R), TGC TTA TTA GCA CAA GAC CTC AAG (609L), ATC ACC CTA TTC CCA GTG GA (850R), GCA GAT GAT AAG CTA TCC TTG AGA (850L), CAT CAC TGA GTC TTG CTG GTT T (1011R), CAA TTT AGG GGA AGG AAG CA (1011L), TGG TAG CTG GGC ATA AAA GC (1560R), AAT GGG AGA AAA GGC AGG AT (1560L), CAG GAA AGG GTG TGT GTG TG (1755R), and TAC GCT CTG GCA GTT TTC AA (1755L). .. The PCR products were gel extracted using the Wizard PCR Preps DNA purification system (Promega).

Electroporation:

Article Title: Mapping of Human Autoantibody Binding Sites on the Calcium-Sensing Receptor
Article Snippet: The DNA fragments were purified using a Wizard PCR Preps DNA Purification System and treated with T4 DNA polymerase (Promega) according to the manufacturer's protocol to create blunt ends. .. After further purification, the fragments were ligated into the Eco RV restriction site of pComb3 using standard methods. ( ) The CaSR cDNA fragment library was recovered by electroporation of Escherichia coli XL1-Blue cells (Stratagene, La Jolla, CA, USA), as described by the manufacturer.

Southern Blot:

Article Title: AgtA, the Dicarboxylic Amino Acid Transporter of Aspergillus nidulans, Is Concertedly Down-Regulated by Exquisite Sensitivity to Nitrogen Metabolite Repression and Ammonium-Elicited Endocytosis ▿, Is Concertedly Down-Regulated by Exquisite Sensitivity to Nitrogen Metabolite Repression and Ammonium-Elicited Endocytosis ▿ †
Article Snippet: Southern blot analysis was carried out according to Southern ( ) and Sambrook and Russell ( ). .. DNA bands were purified from agarose gels with the Wizard PCR Preps DNA purification system (Promega).

Concentration Assay:

Article Title: Differential effects of a common splice site polymorphism on the generation of OAS1 variants in human bronchial epithelial cells.
Article Snippet: Isoform-specific RT-PCR products containing the target sequences, amplified with primers in Supplementary Table 1, were purified using the Wizard PCR Preps DNA Purification System (Promega, Fitchburg, WI, USA). .. Their copy numbers were calculated from the concentration of DNA determined by measuring absorbance at 260 nm.

Nick Translation:

Article Title: A Deletion at the Mouse Xist Gene Exposes Trans-effects That Alter the Heterochromatin of the Inactive X Chromosome and the Replication Time and DNA Stability of Both X Chromosomes
Article Snippet: The PCR products were gel extracted using the Wizard PCR Preps DNA purification system (Promega). .. For fluorescent probe preparations, 1 mg of DNA was direct labeled with either FluorX-dCTP (for the whole BAC probe) or Cy3-dCTP (for the Xi-specific 6.8-kb combination probe), using the Nick Translation kit (Amersham Biosciences, Arlington Heights, IL).

Infection:

Article Title: A New Huanglongbing Species, "Candidatus Liberibacter psyllaurous," Found To Infect Tomato and Potato, Is Vectored by the Psyllid Bactericera cockerelli (Sulc) ▿
Article Snippet: In all cases (of infected psyllids), this yielded a single visible band of ∼2.6 kb in length. .. Amplified DNA was cleaned using the Wizard PCR Preps DNA purification system (Promega, Madison, WI) and directly sequenced in both directions.

Polymerase Chain Reaction:

Article Title: Mapping of Human Autoantibody Binding Sites on the Calcium-Sensing Receptor
Article Snippet: .. The DNA fragments were purified using a Wizard PCR Preps DNA Purification System and treated with T4 DNA polymerase (Promega) according to the manufacturer's protocol to create blunt ends. .. After further purification, the fragments were ligated into the Eco RV restriction site of pComb3 using standard methods. ( ) The CaSR cDNA fragment library was recovered by electroporation of Escherichia coli XL1-Blue cells (Stratagene, La Jolla, CA, USA), as described by the manufacturer.

Article Title: A Deletion at the Mouse Xist Gene Exposes Trans-effects That Alter the Heterochromatin of the Inactive X Chromosome and the Replication Time and DNA Stability of Both X Chromosomes
Article Snippet: .. The PCR products were gel extracted using the Wizard PCR Preps DNA purification system (Promega). .. To mark both the active and the inactive X chromosomes, mouse BAC RP23-298N24 (obtained from Invitrogen) was used to obtain a second probe.

Article Title: Differential effects of a common splice site polymorphism on the generation of OAS1 variants in human bronchial epithelial cells.
Article Snippet: .. Isoform-specific RT-PCR products containing the target sequences, amplified with primers in Supplementary Table 1, were purified using the Wizard PCR Preps DNA Purification System (Promega, Fitchburg, WI, USA). .. Their copy numbers were calculated from the concentration of DNA determined by measuring absorbance at 260 nm.

Article Title: Cloning and identification of NS5ATP2 gene and its spliced variant transactivated by hepatitis C virus non-structural protein 5A
Article Snippet: .. PCR amplifications products were directly purified by using Wizard PCR Preps DNA Purification System (Promega), and subcloned into pEGM-T easy vectors (Promega) to set up the subtractive library[ , ]. .. On the basis of subtractive cDNA library of genes transactivated by NS5A protein of hepatitis C virus, the coding sequence of a new gene, named NS5ATP2, was obtained by bioinformatics methods.

Article Title: Cloning and expression of 3-deoxy-d-manno-oct-2-ulosonic acid α-ketoside hydrolase from oyster hepatopancreas
Article Snippet: .. The following materials were purchased from commercial sources: marker proteins for molecular weight and p I , Sephacryl S-300 HR, Amersham Pharmacia Biotech; Tris, bis-Tris, MU-α-Neu5Ac and other MU-glycosides, Sigma; phenylmethylsulfonyl fluoride, Pierce; Silica Gel-60 thin-layer chromatography (TLC) plates, EMD Chemicals, Darmstadt, Germany; Centricon-10 and Microcon (10 kDa M r cut-off) micro-concentrators, Amicon; E. coli BL21(DE3), E. coli DH5α, S. cerevisiae INVSc1, P. pastoris , CHO-S cell, pUC18, pET, pYES2, pcDNA3.1/ myc -His(A), pPICZ, 5′-Race System for Rapid Amplification of cDNA Ends (Version 2) and LipofectAMINE 2000, Invitrogen; E. coli Origami, Novagen; GeneAmp PCR System 9600, Perkin-Elmer; Wizard PCR Preps DNA Purification System and pGEM-T Easy Vector System, Promega; RNAlater, Ambion; YPD and SD yeast culture media, BD Bioscience; FastTrack 2.0 Kit, BD Smart Race cDNA Amplification Kit and BD Advantage 2 PCR Kit, Clontech; Thermoscript RT-PCR System, Gibco; Oligotex Direct mRNA midi/maxi Kit and Ni-NTA Superflow column, Qiagen; chemically defined natural Re-LPS from S. minnesota and E. coli D31m4, Rd-LPS from S. minnesota R7 LIST Biological Laboratories, Inc; LPS from Neisseria meningitides strains A1 and M992 were gifts of Dr. Chao-Ming Tsai, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland. .. The α-Kdo-cleaving activity was measured by fluorometric analysis according to the procedure of ) as described previously ( ).

Article Title: Molecular Characterization of Diphtheria Toxin Repressor (dtxR) Genes Present in Nontoxigenic Corynebacterium diphtheriae Strains Isolated in the United Kingdom
Article Snippet: .. The amplicons were purified using the Wizard PCR Preps DNA purification system (Promega Corp., Madison, Wis.) as specified by the manufacturer. .. Sequencing was performed on each strand with the primers used in the initial PCR amplification (DtxR 1F and DtxR 4R) and primers DtxR 2F (5′-CTGAGCGTCTGGAACAATCT-3′) and DtxR 3R (5′-ATCCAACACGGATGTCAGCAT-3′).

Article Title: A New Huanglongbing Species, "Candidatus Liberibacter psyllaurous," Found To Infect Tomato and Potato, Is Vectored by the Psyllid Bactericera cockerelli (Sulc) ▿
Article Snippet: .. Amplified DNA was cleaned using the Wizard PCR Preps DNA purification system (Promega, Madison, WI) and directly sequenced in both directions. .. With the 10F and 480R primers on tomato and potato plants, chloroplast DNA was preferentially amplified.

Article Title: Bovine torovirus: sequencing of the structural genes and expression of the nucleocapsid protein of Breda virus.
Article Snippet: .. Cloning and expression The PCR product containing the BRV N gene was purified using the Wizard PCR Preps DNA Purification System (Promega). .. Amplicons were then ligated to a pET28a(+) cloning vector (Novagen, Madison, WI) , and transformed into INVhF% One Shot cells (Invitrogen, Carlsbad, CA) as per the manufacturer's recommendations.

Article Title: AgtA, the Dicarboxylic Amino Acid Transporter of Aspergillus nidulans, Is Concertedly Down-Regulated by Exquisite Sensitivity to Nitrogen Metabolite Repression and Ammonium-Elicited Endocytosis ▿, Is Concertedly Down-Regulated by Exquisite Sensitivity to Nitrogen Metabolite Repression and Ammonium-Elicited Endocytosis ▿ †
Article Snippet: .. DNA bands were purified from agarose gels with the Wizard PCR Preps DNA purification system (Promega). .. The [α-32 P]dCTP-labeled DNA molecules used as gene-specific probes were prepared with the Megaprime DNA labeling systems kit (Amersham Life Science).

Article Title: First Report of Macrolide-Resistant Strains and Description of a Novel Nucleotide Sequence Variation in the P1 Adhesin Gene in Mycoplasma pneumoniae Clinical Strains Isolated in France over 12 Years ▿
Article Snippet: .. As the PCR-RFLP method did not allow determination of the type of one isolate (isolate Mp3896), the PCR products obtained with primers ADH1 and ADH2 and primers ADH3 and ADH4 ( ) were purified with the Wizard PCR preps DNA purification system (Promega). .. Both strands of the PCR products were sequenced with a BigDye Terminator V1.1 cycle sequencing kit (Applied Biosystems) and a 3130 genetic analyzer (Applied Biosystems/Hitachi), according to the manufacturer's instructions, using primers ADH1, ADH2, ADH3, and ADH4 as well as internal primers.

Article Title: Age-Dependent Demise of GNAS-Mutated Skeletal Stem Cells and "Normalization" of Fibrous Dysplasia of Bone
Article Snippet: .. The 325-bp PCR products were purified using the Promega Wizard PCR Preps DNA Purification System (Madison, WI, USA) and sequenced using a Perkin Elmer Applied Biosystem 377 Automated Sequencer, and the sequence was read from the 3′-5′ (reverse) strand. ..

Article Title: Prevalence and distribution of dermatophytosis lesions on cattle in Plateau State, Nigeria
Article Snippet: .. The amplified DNA was purified using Wizard PCR Preps DNA Purification System (Promega) and sequenced with the primers ITS-1 (5’-TCCGTAGGTGAACCTGCGG-3’) and ITS-4 (5’-TCCTCCGCTTATTGATATGC-3’) using the dye terminator method (Applied Biosystems 377 automatic sequencer). ..

DNA Sequencing:

Article Title: Bovine torovirus: sequencing of the structural genes and expression of the nucleocapsid protein of Breda virus.
Article Snippet: Cloning and expression The PCR product containing the BRV N gene was purified using the Wizard PCR Preps DNA Purification System (Promega). .. Plasmids containing the PCR insert were purified from LB broth cultures using the QIAGEN Plasmid Mini Kit (QIAGEN, Chatsworth, CA), and sequenced using the fmol DNA sequencing system (Promega) as described above.

Article Title: First Report of Macrolide-Resistant Strains and Description of a Novel Nucleotide Sequence Variation in the P1 Adhesin Gene in Mycoplasma pneumoniae Clinical Strains Isolated in France over 12 Years ▿
Article Snippet: Paragraph title: DNA sequencing of the P1 gene. ... As the PCR-RFLP method did not allow determination of the type of one isolate (isolate Mp3896), the PCR products obtained with primers ADH1 and ADH2 and primers ADH3 and ADH4 ( ) were purified with the Wizard PCR preps DNA purification system (Promega).

Article Title: Age-Dependent Demise of GNAS-Mutated Skeletal Stem Cells and "Normalization" of Fibrous Dysplasia of Bone
Article Snippet: A peptide nucleic acid (PNA)-based assay and direct DNA sequencing of the relevant PCR-amplified target sequence in the GNAS gene were used in these experiments to detect the GNAS mutations in the different populations of bone marrow stromal cells (BMSCs) as described previously ( ). ( ) Direct DNA sequencing can detect only one mutant cell out of two-three cells. .. The 325-bp PCR products were purified using the Promega Wizard PCR Preps DNA Purification System (Madison, WI, USA) and sequenced using a Perkin Elmer Applied Biosystem 377 Automated Sequencer, and the sequence was read from the 3′-5′ (reverse) strand.

DNA Labeling:

Article Title: AgtA, the Dicarboxylic Amino Acid Transporter of Aspergillus nidulans, Is Concertedly Down-Regulated by Exquisite Sensitivity to Nitrogen Metabolite Repression and Ammonium-Elicited Endocytosis ▿, Is Concertedly Down-Regulated by Exquisite Sensitivity to Nitrogen Metabolite Repression and Ammonium-Elicited Endocytosis ▿ †
Article Snippet: DNA bands were purified from agarose gels with the Wizard PCR Preps DNA purification system (Promega). .. The [α-32 P]dCTP-labeled DNA molecules used as gene-specific probes were prepared with the Megaprime DNA labeling systems kit (Amersham Life Science).

Reverse Transcription Polymerase Chain Reaction:

Article Title: Differential effects of a common splice site polymorphism on the generation of OAS1 variants in human bronchial epithelial cells.
Article Snippet: .. Isoform-specific RT-PCR products containing the target sequences, amplified with primers in Supplementary Table 1, were purified using the Wizard PCR Preps DNA Purification System (Promega, Fitchburg, WI, USA). .. Their copy numbers were calculated from the concentration of DNA determined by measuring absorbance at 260 nm.

Article Title: Cloning and expression of 3-deoxy-d-manno-oct-2-ulosonic acid α-ketoside hydrolase from oyster hepatopancreas
Article Snippet: .. The following materials were purchased from commercial sources: marker proteins for molecular weight and p I , Sephacryl S-300 HR, Amersham Pharmacia Biotech; Tris, bis-Tris, MU-α-Neu5Ac and other MU-glycosides, Sigma; phenylmethylsulfonyl fluoride, Pierce; Silica Gel-60 thin-layer chromatography (TLC) plates, EMD Chemicals, Darmstadt, Germany; Centricon-10 and Microcon (10 kDa M r cut-off) micro-concentrators, Amicon; E. coli BL21(DE3), E. coli DH5α, S. cerevisiae INVSc1, P. pastoris , CHO-S cell, pUC18, pET, pYES2, pcDNA3.1/ myc -His(A), pPICZ, 5′-Race System for Rapid Amplification of cDNA Ends (Version 2) and LipofectAMINE 2000, Invitrogen; E. coli Origami, Novagen; GeneAmp PCR System 9600, Perkin-Elmer; Wizard PCR Preps DNA Purification System and pGEM-T Easy Vector System, Promega; RNAlater, Ambion; YPD and SD yeast culture media, BD Bioscience; FastTrack 2.0 Kit, BD Smart Race cDNA Amplification Kit and BD Advantage 2 PCR Kit, Clontech; Thermoscript RT-PCR System, Gibco; Oligotex Direct mRNA midi/maxi Kit and Ni-NTA Superflow column, Qiagen; chemically defined natural Re-LPS from S. minnesota and E. coli D31m4, Rd-LPS from S. minnesota R7 LIST Biological Laboratories, Inc; LPS from Neisseria meningitides strains A1 and M992 were gifts of Dr. Chao-Ming Tsai, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland. .. The α-Kdo-cleaving activity was measured by fluorometric analysis according to the procedure of ) as described previously ( ).

Binding Assay:

Article Title: Age-Dependent Demise of GNAS-Mutated Skeletal Stem Cells and "Normalization" of Fibrous Dysplasia of Bone
Article Snippet: Consequently, a peptide nucleic acid (PNA) sequence complementary to the wildtype sequence was added to the reaction mixture to prevent the binding of the sense primer to the normal allele. .. The 325-bp PCR products were purified using the Promega Wizard PCR Preps DNA Purification System (Madison, WI, USA) and sequenced using a Perkin Elmer Applied Biosystem 377 Automated Sequencer, and the sequence was read from the 3′-5′ (reverse) strand.

Cellular Antioxidant Activity Assay:

Article Title: A Deletion at the Mouse Xist Gene Exposes Trans-effects That Alter the Heterochromatin of the Inactive X Chromosome and the Replication Time and DNA Stability of Both X Chromosomes
Article Snippet: The sequences of primers are: GCA AGC ATA AGG ACC AGA GC (412R), TTC CAC AAG AAA TAT TAC ACA AAA CA (412L), CCT AAC CAT TGA GCC GTC TT (605R), GGT CTC TGA ACT ACC AAT TGC AC (605L), GCA ATG ACA AAT GTT TTG TGG (609R), TGC TTA TTA GCA CAA GAC CTC AAG (609L), ATC ACC CTA TTC CCA GTG GA (850R), GCA GAT GAT AAG CTA TCC TTG AGA (850L), CAT CAC TGA GTC TTG CTG GTT T (1011R), CAA TTT AGG GGA AGG AAG CA (1011L), TGG TAG CTG GGC ATA AAA GC (1560R), AAT GGG AGA AAA GGC AGG AT (1560L), CAG GAA AGG GTG TGT GTG TG (1755R), and TAC GCT CTG GCA GTT TTC AA (1755L). .. The PCR products were gel extracted using the Wizard PCR Preps DNA purification system (Promega).

Molecular Weight:

Article Title: Cloning and expression of 3-deoxy-d-manno-oct-2-ulosonic acid α-ketoside hydrolase from oyster hepatopancreas
Article Snippet: .. The following materials were purchased from commercial sources: marker proteins for molecular weight and p I , Sephacryl S-300 HR, Amersham Pharmacia Biotech; Tris, bis-Tris, MU-α-Neu5Ac and other MU-glycosides, Sigma; phenylmethylsulfonyl fluoride, Pierce; Silica Gel-60 thin-layer chromatography (TLC) plates, EMD Chemicals, Darmstadt, Germany; Centricon-10 and Microcon (10 kDa M r cut-off) micro-concentrators, Amicon; E. coli BL21(DE3), E. coli DH5α, S. cerevisiae INVSc1, P. pastoris , CHO-S cell, pUC18, pET, pYES2, pcDNA3.1/ myc -His(A), pPICZ, 5′-Race System for Rapid Amplification of cDNA Ends (Version 2) and LipofectAMINE 2000, Invitrogen; E. coli Origami, Novagen; GeneAmp PCR System 9600, Perkin-Elmer; Wizard PCR Preps DNA Purification System and pGEM-T Easy Vector System, Promega; RNAlater, Ambion; YPD and SD yeast culture media, BD Bioscience; FastTrack 2.0 Kit, BD Smart Race cDNA Amplification Kit and BD Advantage 2 PCR Kit, Clontech; Thermoscript RT-PCR System, Gibco; Oligotex Direct mRNA midi/maxi Kit and Ni-NTA Superflow column, Qiagen; chemically defined natural Re-LPS from S. minnesota and E. coli D31m4, Rd-LPS from S. minnesota R7 LIST Biological Laboratories, Inc; LPS from Neisseria meningitides strains A1 and M992 were gifts of Dr. Chao-Ming Tsai, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland. .. The α-Kdo-cleaving activity was measured by fluorometric analysis according to the procedure of ) as described previously ( ).

DNA Purification:

Article Title: Mapping of Human Autoantibody Binding Sites on the Calcium-Sensing Receptor
Article Snippet: .. The DNA fragments were purified using a Wizard PCR Preps DNA Purification System and treated with T4 DNA polymerase (Promega) according to the manufacturer's protocol to create blunt ends. .. After further purification, the fragments were ligated into the Eco RV restriction site of pComb3 using standard methods. ( ) The CaSR cDNA fragment library was recovered by electroporation of Escherichia coli XL1-Blue cells (Stratagene, La Jolla, CA, USA), as described by the manufacturer.

Article Title: A Deletion at the Mouse Xist Gene Exposes Trans-effects That Alter the Heterochromatin of the Inactive X Chromosome and the Replication Time and DNA Stability of Both X Chromosomes
Article Snippet: .. The PCR products were gel extracted using the Wizard PCR Preps DNA purification system (Promega). .. To mark both the active and the inactive X chromosomes, mouse BAC RP23-298N24 (obtained from Invitrogen) was used to obtain a second probe.

Article Title: Differential effects of a common splice site polymorphism on the generation of OAS1 variants in human bronchial epithelial cells.
Article Snippet: .. Isoform-specific RT-PCR products containing the target sequences, amplified with primers in Supplementary Table 1, were purified using the Wizard PCR Preps DNA Purification System (Promega, Fitchburg, WI, USA). .. Their copy numbers were calculated from the concentration of DNA determined by measuring absorbance at 260 nm.

Article Title: Cloning and identification of NS5ATP2 gene and its spliced variant transactivated by hepatitis C virus non-structural protein 5A
Article Snippet: .. PCR amplifications products were directly purified by using Wizard PCR Preps DNA Purification System (Promega), and subcloned into pEGM-T easy vectors (Promega) to set up the subtractive library[ , ]. .. On the basis of subtractive cDNA library of genes transactivated by NS5A protein of hepatitis C virus, the coding sequence of a new gene, named NS5ATP2, was obtained by bioinformatics methods.

Article Title: Cloning and expression of 3-deoxy-d-manno-oct-2-ulosonic acid α-ketoside hydrolase from oyster hepatopancreas
Article Snippet: .. The following materials were purchased from commercial sources: marker proteins for molecular weight and p I , Sephacryl S-300 HR, Amersham Pharmacia Biotech; Tris, bis-Tris, MU-α-Neu5Ac and other MU-glycosides, Sigma; phenylmethylsulfonyl fluoride, Pierce; Silica Gel-60 thin-layer chromatography (TLC) plates, EMD Chemicals, Darmstadt, Germany; Centricon-10 and Microcon (10 kDa M r cut-off) micro-concentrators, Amicon; E. coli BL21(DE3), E. coli DH5α, S. cerevisiae INVSc1, P. pastoris , CHO-S cell, pUC18, pET, pYES2, pcDNA3.1/ myc -His(A), pPICZ, 5′-Race System for Rapid Amplification of cDNA Ends (Version 2) and LipofectAMINE 2000, Invitrogen; E. coli Origami, Novagen; GeneAmp PCR System 9600, Perkin-Elmer; Wizard PCR Preps DNA Purification System and pGEM-T Easy Vector System, Promega; RNAlater, Ambion; YPD and SD yeast culture media, BD Bioscience; FastTrack 2.0 Kit, BD Smart Race cDNA Amplification Kit and BD Advantage 2 PCR Kit, Clontech; Thermoscript RT-PCR System, Gibco; Oligotex Direct mRNA midi/maxi Kit and Ni-NTA Superflow column, Qiagen; chemically defined natural Re-LPS from S. minnesota and E. coli D31m4, Rd-LPS from S. minnesota R7 LIST Biological Laboratories, Inc; LPS from Neisseria meningitides strains A1 and M992 were gifts of Dr. Chao-Ming Tsai, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland. .. The α-Kdo-cleaving activity was measured by fluorometric analysis according to the procedure of ) as described previously ( ).

Article Title: Molecular Characterization of Diphtheria Toxin Repressor (dtxR) Genes Present in Nontoxigenic Corynebacterium diphtheriae Strains Isolated in the United Kingdom
Article Snippet: .. The amplicons were purified using the Wizard PCR Preps DNA purification system (Promega Corp., Madison, Wis.) as specified by the manufacturer. .. Sequencing was performed on each strand with the primers used in the initial PCR amplification (DtxR 1F and DtxR 4R) and primers DtxR 2F (5′-CTGAGCGTCTGGAACAATCT-3′) and DtxR 3R (5′-ATCCAACACGGATGTCAGCAT-3′).

Article Title: A New Huanglongbing Species, "Candidatus Liberibacter psyllaurous," Found To Infect Tomato and Potato, Is Vectored by the Psyllid Bactericera cockerelli (Sulc) ▿
Article Snippet: .. Amplified DNA was cleaned using the Wizard PCR Preps DNA purification system (Promega, Madison, WI) and directly sequenced in both directions. .. With the 10F and 480R primers on tomato and potato plants, chloroplast DNA was preferentially amplified.

Article Title: Bovine torovirus: sequencing of the structural genes and expression of the nucleocapsid protein of Breda virus.
Article Snippet: .. Cloning and expression The PCR product containing the BRV N gene was purified using the Wizard PCR Preps DNA Purification System (Promega). .. Amplicons were then ligated to a pET28a(+) cloning vector (Novagen, Madison, WI) , and transformed into INVhF% One Shot cells (Invitrogen, Carlsbad, CA) as per the manufacturer's recommendations.

Article Title: AgtA, the Dicarboxylic Amino Acid Transporter of Aspergillus nidulans, Is Concertedly Down-Regulated by Exquisite Sensitivity to Nitrogen Metabolite Repression and Ammonium-Elicited Endocytosis ▿, Is Concertedly Down-Regulated by Exquisite Sensitivity to Nitrogen Metabolite Repression and Ammonium-Elicited Endocytosis ▿ †
Article Snippet: .. DNA bands were purified from agarose gels with the Wizard PCR Preps DNA purification system (Promega). .. The [α-32 P]dCTP-labeled DNA molecules used as gene-specific probes were prepared with the Megaprime DNA labeling systems kit (Amersham Life Science).

Article Title: First Report of Macrolide-Resistant Strains and Description of a Novel Nucleotide Sequence Variation in the P1 Adhesin Gene in Mycoplasma pneumoniae Clinical Strains Isolated in France over 12 Years ▿
Article Snippet: .. As the PCR-RFLP method did not allow determination of the type of one isolate (isolate Mp3896), the PCR products obtained with primers ADH1 and ADH2 and primers ADH3 and ADH4 ( ) were purified with the Wizard PCR preps DNA purification system (Promega). .. Both strands of the PCR products were sequenced with a BigDye Terminator V1.1 cycle sequencing kit (Applied Biosystems) and a 3130 genetic analyzer (Applied Biosystems/Hitachi), according to the manufacturer's instructions, using primers ADH1, ADH2, ADH3, and ADH4 as well as internal primers.

Article Title: Age-Dependent Demise of GNAS-Mutated Skeletal Stem Cells and "Normalization" of Fibrous Dysplasia of Bone
Article Snippet: .. The 325-bp PCR products were purified using the Promega Wizard PCR Preps DNA Purification System (Madison, WI, USA) and sequenced using a Perkin Elmer Applied Biosystem 377 Automated Sequencer, and the sequence was read from the 3′-5′ (reverse) strand. ..

Article Title: Prevalence and distribution of dermatophytosis lesions on cattle in Plateau State, Nigeria
Article Snippet: .. The amplified DNA was purified using Wizard PCR Preps DNA Purification System (Promega) and sequenced with the primers ITS-1 (5’-TCCGTAGGTGAACCTGCGG-3’) and ITS-4 (5’-TCCTCCGCTTATTGATATGC-3’) using the dye terminator method (Applied Biosystems 377 automatic sequencer). ..

Mutagenesis:

Article Title: Age-Dependent Demise of GNAS-Mutated Skeletal Stem Cells and "Normalization" of Fibrous Dysplasia of Bone
Article Snippet: Paragraph title: Mutation analysis ... The 325-bp PCR products were purified using the Promega Wizard PCR Preps DNA Purification System (Madison, WI, USA) and sequenced using a Perkin Elmer Applied Biosystem 377 Automated Sequencer, and the sequence was read from the 3′-5′ (reverse) strand.

Isolation:

Article Title: Age-Dependent Demise of GNAS-Mutated Skeletal Stem Cells and "Normalization" of Fibrous Dysplasia of Bone
Article Snippet: Genomic DNA was isolated from cells using the DNeasy Tissue Kit according to the manufacturer’s instructions (Qiagen, Valencia, CA, USA). .. The 325-bp PCR products were purified using the Promega Wizard PCR Preps DNA Purification System (Madison, WI, USA) and sequenced using a Perkin Elmer Applied Biosystem 377 Automated Sequencer, and the sequence was read from the 3′-5′ (reverse) strand.

Labeling:

Article Title: A Deletion at the Mouse Xist Gene Exposes Trans-effects That Alter the Heterochromatin of the Inactive X Chromosome and the Replication Time and DNA Stability of Both X Chromosomes
Article Snippet: The PCR products were gel extracted using the Wizard PCR Preps DNA purification system (Promega). .. For fluorescent probe preparations, 1 mg of DNA was direct labeled with either FluorX-dCTP (for the whole BAC probe) or Cy3-dCTP (for the Xi-specific 6.8-kb combination probe), using the Nick Translation kit (Amersham Biosciences, Arlington Heights, IL).

Purification:

Article Title: Mapping of Human Autoantibody Binding Sites on the Calcium-Sensing Receptor
Article Snippet: .. The DNA fragments were purified using a Wizard PCR Preps DNA Purification System and treated with T4 DNA polymerase (Promega) according to the manufacturer's protocol to create blunt ends. .. After further purification, the fragments were ligated into the Eco RV restriction site of pComb3 using standard methods. ( ) The CaSR cDNA fragment library was recovered by electroporation of Escherichia coli XL1-Blue cells (Stratagene, La Jolla, CA, USA), as described by the manufacturer.

Article Title: Differential effects of a common splice site polymorphism on the generation of OAS1 variants in human bronchial epithelial cells.
Article Snippet: .. Isoform-specific RT-PCR products containing the target sequences, amplified with primers in Supplementary Table 1, were purified using the Wizard PCR Preps DNA Purification System (Promega, Fitchburg, WI, USA). .. Their copy numbers were calculated from the concentration of DNA determined by measuring absorbance at 260 nm.

Article Title: Cloning and identification of NS5ATP2 gene and its spliced variant transactivated by hepatitis C virus non-structural protein 5A
Article Snippet: .. PCR amplifications products were directly purified by using Wizard PCR Preps DNA Purification System (Promega), and subcloned into pEGM-T easy vectors (Promega) to set up the subtractive library[ , ]. .. On the basis of subtractive cDNA library of genes transactivated by NS5A protein of hepatitis C virus, the coding sequence of a new gene, named NS5ATP2, was obtained by bioinformatics methods.

Article Title: Molecular Characterization of Diphtheria Toxin Repressor (dtxR) Genes Present in Nontoxigenic Corynebacterium diphtheriae Strains Isolated in the United Kingdom
Article Snippet: .. The amplicons were purified using the Wizard PCR Preps DNA purification system (Promega Corp., Madison, Wis.) as specified by the manufacturer. .. Sequencing was performed on each strand with the primers used in the initial PCR amplification (DtxR 1F and DtxR 4R) and primers DtxR 2F (5′-CTGAGCGTCTGGAACAATCT-3′) and DtxR 3R (5′-ATCCAACACGGATGTCAGCAT-3′).

Article Title: Bovine torovirus: sequencing of the structural genes and expression of the nucleocapsid protein of Breda virus.
Article Snippet: .. Cloning and expression The PCR product containing the BRV N gene was purified using the Wizard PCR Preps DNA Purification System (Promega). .. Amplicons were then ligated to a pET28a(+) cloning vector (Novagen, Madison, WI) , and transformed into INVhF% One Shot cells (Invitrogen, Carlsbad, CA) as per the manufacturer's recommendations.

Article Title: AgtA, the Dicarboxylic Amino Acid Transporter of Aspergillus nidulans, Is Concertedly Down-Regulated by Exquisite Sensitivity to Nitrogen Metabolite Repression and Ammonium-Elicited Endocytosis ▿, Is Concertedly Down-Regulated by Exquisite Sensitivity to Nitrogen Metabolite Repression and Ammonium-Elicited Endocytosis ▿ †
Article Snippet: .. DNA bands were purified from agarose gels with the Wizard PCR Preps DNA purification system (Promega). .. The [α-32 P]dCTP-labeled DNA molecules used as gene-specific probes were prepared with the Megaprime DNA labeling systems kit (Amersham Life Science).

Article Title: First Report of Macrolide-Resistant Strains and Description of a Novel Nucleotide Sequence Variation in the P1 Adhesin Gene in Mycoplasma pneumoniae Clinical Strains Isolated in France over 12 Years ▿
Article Snippet: .. As the PCR-RFLP method did not allow determination of the type of one isolate (isolate Mp3896), the PCR products obtained with primers ADH1 and ADH2 and primers ADH3 and ADH4 ( ) were purified with the Wizard PCR preps DNA purification system (Promega). .. Both strands of the PCR products were sequenced with a BigDye Terminator V1.1 cycle sequencing kit (Applied Biosystems) and a 3130 genetic analyzer (Applied Biosystems/Hitachi), according to the manufacturer's instructions, using primers ADH1, ADH2, ADH3, and ADH4 as well as internal primers.

Article Title: Age-Dependent Demise of GNAS-Mutated Skeletal Stem Cells and "Normalization" of Fibrous Dysplasia of Bone
Article Snippet: .. The 325-bp PCR products were purified using the Promega Wizard PCR Preps DNA Purification System (Madison, WI, USA) and sequenced using a Perkin Elmer Applied Biosystem 377 Automated Sequencer, and the sequence was read from the 3′-5′ (reverse) strand. ..

Article Title: Prevalence and distribution of dermatophytosis lesions on cattle in Plateau State, Nigeria
Article Snippet: .. The amplified DNA was purified using Wizard PCR Preps DNA Purification System (Promega) and sequenced with the primers ITS-1 (5’-TCCGTAGGTGAACCTGCGG-3’) and ITS-4 (5’-TCCTCCGCTTATTGATATGC-3’) using the dye terminator method (Applied Biosystems 377 automatic sequencer). ..

Sequencing:

Article Title: Molecular Characterization of Diphtheria Toxin Repressor (dtxR) Genes Present in Nontoxigenic Corynebacterium diphtheriae Strains Isolated in the United Kingdom
Article Snippet: Paragraph title: dtxR gene sequencing. ... The amplicons were purified using the Wizard PCR Preps DNA purification system (Promega Corp., Madison, Wis.) as specified by the manufacturer.

Article Title: A New Huanglongbing Species, "Candidatus Liberibacter psyllaurous," Found To Infect Tomato and Potato, Is Vectored by the Psyllid Bactericera cockerelli (Sulc) ▿
Article Snippet: Amplified DNA was cleaned using the Wizard PCR Preps DNA purification system (Promega, Madison, WI) and directly sequenced in both directions. .. Consequently, the four overlapping sets of primers designed by primer walking (see above and Table ) were used to amplify and subsequently sequence HLB bacteria DNA from plants.

Article Title: First Report of Macrolide-Resistant Strains and Description of a Novel Nucleotide Sequence Variation in the P1 Adhesin Gene in Mycoplasma pneumoniae Clinical Strains Isolated in France over 12 Years ▿
Article Snippet: As the PCR-RFLP method did not allow determination of the type of one isolate (isolate Mp3896), the PCR products obtained with primers ADH1 and ADH2 and primers ADH3 and ADH4 ( ) were purified with the Wizard PCR preps DNA purification system (Promega). .. Both strands of the PCR products were sequenced with a BigDye Terminator V1.1 cycle sequencing kit (Applied Biosystems) and a 3130 genetic analyzer (Applied Biosystems/Hitachi), according to the manufacturer's instructions, using primers ADH1, ADH2, ADH3, and ADH4 as well as internal primers.

Article Title: Age-Dependent Demise of GNAS-Mutated Skeletal Stem Cells and "Normalization" of Fibrous Dysplasia of Bone
Article Snippet: .. The 325-bp PCR products were purified using the Promega Wizard PCR Preps DNA Purification System (Madison, WI, USA) and sequenced using a Perkin Elmer Applied Biosystem 377 Automated Sequencer, and the sequence was read from the 3′-5′ (reverse) strand. ..

Blocking Assay:

Article Title: Age-Dependent Demise of GNAS-Mutated Skeletal Stem Cells and "Normalization" of Fibrous Dysplasia of Bone
Article Snippet: The samples were heated to 94°C for 15 min, cycled 40 times (94°C for 30 s, 68°C for 60 s [to allow the PNA to bind specifically to any nonmutant allele and block the annealing of the forward primer], 55°C for 30 s, and 72°C for 60 s), and terminated for 7 min at 72°C. .. The 325-bp PCR products were purified using the Promega Wizard PCR Preps DNA Purification System (Madison, WI, USA) and sequenced using a Perkin Elmer Applied Biosystem 377 Automated Sequencer, and the sequence was read from the 3′-5′ (reverse) strand.

Activated Clotting Time Assay:

Article Title: A Deletion at the Mouse Xist Gene Exposes Trans-effects That Alter the Heterochromatin of the Inactive X Chromosome and the Replication Time and DNA Stability of Both X Chromosomes
Article Snippet: The sequences of primers are: GCA AGC ATA AGG ACC AGA GC (412R), TTC CAC AAG AAA TAT TAC ACA AAA CA (412L), CCT AAC CAT TGA GCC GTC TT (605R), GGT CTC TGA ACT ACC AAT TGC AC (605L), GCA ATG ACA AAT GTT TTG TGG (609R), TGC TTA TTA GCA CAA GAC CTC AAG (609L), ATC ACC CTA TTC CCA GTG GA (850R), GCA GAT GAT AAG CTA TCC TTG AGA (850L), CAT CAC TGA GTC TTG CTG GTT T (1011R), CAA TTT AGG GGA AGG AAG CA (1011L), TGG TAG CTG GGC ATA AAA GC (1560R), AAT GGG AGA AAA GGC AGG AT (1560L), CAG GAA AGG GTG TGT GTG TG (1755R), and TAC GCT CTG GCA GTT TTC AA (1755L). .. The PCR products were gel extracted using the Wizard PCR Preps DNA purification system (Promega).

Rapid Amplification of cDNA Ends:

Article Title: Cloning and expression of 3-deoxy-d-manno-oct-2-ulosonic acid α-ketoside hydrolase from oyster hepatopancreas
Article Snippet: .. The following materials were purchased from commercial sources: marker proteins for molecular weight and p I , Sephacryl S-300 HR, Amersham Pharmacia Biotech; Tris, bis-Tris, MU-α-Neu5Ac and other MU-glycosides, Sigma; phenylmethylsulfonyl fluoride, Pierce; Silica Gel-60 thin-layer chromatography (TLC) plates, EMD Chemicals, Darmstadt, Germany; Centricon-10 and Microcon (10 kDa M r cut-off) micro-concentrators, Amicon; E. coli BL21(DE3), E. coli DH5α, S. cerevisiae INVSc1, P. pastoris , CHO-S cell, pUC18, pET, pYES2, pcDNA3.1/ myc -His(A), pPICZ, 5′-Race System for Rapid Amplification of cDNA Ends (Version 2) and LipofectAMINE 2000, Invitrogen; E. coli Origami, Novagen; GeneAmp PCR System 9600, Perkin-Elmer; Wizard PCR Preps DNA Purification System and pGEM-T Easy Vector System, Promega; RNAlater, Ambion; YPD and SD yeast culture media, BD Bioscience; FastTrack 2.0 Kit, BD Smart Race cDNA Amplification Kit and BD Advantage 2 PCR Kit, Clontech; Thermoscript RT-PCR System, Gibco; Oligotex Direct mRNA midi/maxi Kit and Ni-NTA Superflow column, Qiagen; chemically defined natural Re-LPS from S. minnesota and E. coli D31m4, Rd-LPS from S. minnesota R7 LIST Biological Laboratories, Inc; LPS from Neisseria meningitides strains A1 and M992 were gifts of Dr. Chao-Ming Tsai, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland. .. The α-Kdo-cleaving activity was measured by fluorometric analysis according to the procedure of ) as described previously ( ).

Plasmid Preparation:

Article Title: Mapping of Human Autoantibody Binding Sites on the Calcium-Sensing Receptor
Article Snippet: To construct a CaSR cDNA fragment library in pComb3, full-length CaSR cDNA was prepared from pcCaSR-FLAG( ) by restriction of the plasmid with Kpn I (Promega) and Xba I (Promega). .. The DNA fragments were purified using a Wizard PCR Preps DNA Purification System and treated with T4 DNA polymerase (Promega) according to the manufacturer's protocol to create blunt ends.

Article Title: Cloning and expression of 3-deoxy-d-manno-oct-2-ulosonic acid α-ketoside hydrolase from oyster hepatopancreas
Article Snippet: .. The following materials were purchased from commercial sources: marker proteins for molecular weight and p I , Sephacryl S-300 HR, Amersham Pharmacia Biotech; Tris, bis-Tris, MU-α-Neu5Ac and other MU-glycosides, Sigma; phenylmethylsulfonyl fluoride, Pierce; Silica Gel-60 thin-layer chromatography (TLC) plates, EMD Chemicals, Darmstadt, Germany; Centricon-10 and Microcon (10 kDa M r cut-off) micro-concentrators, Amicon; E. coli BL21(DE3), E. coli DH5α, S. cerevisiae INVSc1, P. pastoris , CHO-S cell, pUC18, pET, pYES2, pcDNA3.1/ myc -His(A), pPICZ, 5′-Race System for Rapid Amplification of cDNA Ends (Version 2) and LipofectAMINE 2000, Invitrogen; E. coli Origami, Novagen; GeneAmp PCR System 9600, Perkin-Elmer; Wizard PCR Preps DNA Purification System and pGEM-T Easy Vector System, Promega; RNAlater, Ambion; YPD and SD yeast culture media, BD Bioscience; FastTrack 2.0 Kit, BD Smart Race cDNA Amplification Kit and BD Advantage 2 PCR Kit, Clontech; Thermoscript RT-PCR System, Gibco; Oligotex Direct mRNA midi/maxi Kit and Ni-NTA Superflow column, Qiagen; chemically defined natural Re-LPS from S. minnesota and E. coli D31m4, Rd-LPS from S. minnesota R7 LIST Biological Laboratories, Inc; LPS from Neisseria meningitides strains A1 and M992 were gifts of Dr. Chao-Ming Tsai, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland. .. The α-Kdo-cleaving activity was measured by fluorometric analysis according to the procedure of ) as described previously ( ).

Article Title: Bovine torovirus: sequencing of the structural genes and expression of the nucleocapsid protein of Breda virus.
Article Snippet: Cloning and expression The PCR product containing the BRV N gene was purified using the Wizard PCR Preps DNA Purification System (Promega). .. Amplicons were then ligated to a pET28a(+) cloning vector (Novagen, Madison, WI) , and transformed into INVhF% One Shot cells (Invitrogen, Carlsbad, CA) as per the manufacturer's recommendations.

Article Title: AgtA, the Dicarboxylic Amino Acid Transporter of Aspergillus nidulans, Is Concertedly Down-Regulated by Exquisite Sensitivity to Nitrogen Metabolite Repression and Ammonium-Elicited Endocytosis ▿, Is Concertedly Down-Regulated by Exquisite Sensitivity to Nitrogen Metabolite Repression and Ammonium-Elicited Endocytosis ▿ †
Article Snippet: TA cloning was carried out with the pGEM-T Easy vector system (Promega). .. DNA bands were purified from agarose gels with the Wizard PCR Preps DNA purification system (Promega).

Software:

Article Title: Molecular Characterization of Diphtheria Toxin Repressor (dtxR) Genes Present in Nontoxigenic Corynebacterium diphtheriae Strains Isolated in the United Kingdom
Article Snippet: The amplicons were purified using the Wizard PCR Preps DNA purification system (Promega Corp., Madison, Wis.) as specified by the manufacturer. .. Raw chromatograph data were edited using Genebuilder sequence analysis software, and multiple alignments were made in CLUSTAL W , which is part of the BioEdit software package ( ).

RNA Extraction:

Article Title: Cloning and identification of NS5ATP2 gene and its spliced variant transactivated by hepatitis C virus non-structural protein 5A
Article Snippet: Paragraph title: RNA extraction and SSH ... PCR amplifications products were directly purified by using Wizard PCR Preps DNA Purification System (Promega), and subcloned into pEGM-T easy vectors (Promega) to set up the subtractive library[ , ].

Positron Emission Tomography:

Article Title: Cloning and expression of 3-deoxy-d-manno-oct-2-ulosonic acid α-ketoside hydrolase from oyster hepatopancreas
Article Snippet: .. The following materials were purchased from commercial sources: marker proteins for molecular weight and p I , Sephacryl S-300 HR, Amersham Pharmacia Biotech; Tris, bis-Tris, MU-α-Neu5Ac and other MU-glycosides, Sigma; phenylmethylsulfonyl fluoride, Pierce; Silica Gel-60 thin-layer chromatography (TLC) plates, EMD Chemicals, Darmstadt, Germany; Centricon-10 and Microcon (10 kDa M r cut-off) micro-concentrators, Amicon; E. coli BL21(DE3), E. coli DH5α, S. cerevisiae INVSc1, P. pastoris , CHO-S cell, pUC18, pET, pYES2, pcDNA3.1/ myc -His(A), pPICZ, 5′-Race System for Rapid Amplification of cDNA Ends (Version 2) and LipofectAMINE 2000, Invitrogen; E. coli Origami, Novagen; GeneAmp PCR System 9600, Perkin-Elmer; Wizard PCR Preps DNA Purification System and pGEM-T Easy Vector System, Promega; RNAlater, Ambion; YPD and SD yeast culture media, BD Bioscience; FastTrack 2.0 Kit, BD Smart Race cDNA Amplification Kit and BD Advantage 2 PCR Kit, Clontech; Thermoscript RT-PCR System, Gibco; Oligotex Direct mRNA midi/maxi Kit and Ni-NTA Superflow column, Qiagen; chemically defined natural Re-LPS from S. minnesota and E. coli D31m4, Rd-LPS from S. minnesota R7 LIST Biological Laboratories, Inc; LPS from Neisseria meningitides strains A1 and M992 were gifts of Dr. Chao-Ming Tsai, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland. .. The α-Kdo-cleaving activity was measured by fluorometric analysis according to the procedure of ) as described previously ( ).

Agarose Gel Electrophoresis:

Article Title: Mapping of Human Autoantibody Binding Sites on the Calcium-Sensing Receptor
Article Snippet: The resulting 3255-bp CaSR cDNA fragment was separated by agarose gel electrophoresis( ) and purified using a Wizard PCR Preps DNA Purification System (Promega, Southampton, UK). .. The DNA fragments were purified using a Wizard PCR Preps DNA Purification System and treated with T4 DNA polymerase (Promega) according to the manufacturer's protocol to create blunt ends.

Article Title: Differential effects of a common splice site polymorphism on the generation of OAS1 variants in human bronchial epithelial cells.
Article Snippet: Specific amplification of the target was confirmed by a single peak in the dissociation curve and by visualization of the expected size of RT-PCR products on an agarose gel. .. Isoform-specific RT-PCR products containing the target sequences, amplified with primers in Supplementary Table 1, were purified using the Wizard PCR Preps DNA Purification System (Promega, Fitchburg, WI, USA).

Article Title: A New Huanglongbing Species, "Candidatus Liberibacter psyllaurous," Found To Infect Tomato and Potato, Is Vectored by the Psyllid Bactericera cockerelli (Sulc) ▿
Article Snippet: To ensure that one bacterial symbiont was present ( , ), 10F and 480R PCR products of an HLB-infected psyllid individual (from the Texas colony [see below]) were run out on a 0.7% agarose gel at ∼1.7 V/cm. .. Amplified DNA was cleaned using the Wizard PCR Preps DNA purification system (Promega, Madison, WI) and directly sequenced in both directions.

Chromosome Walking:

Article Title: A New Huanglongbing Species, "Candidatus Liberibacter psyllaurous," Found To Infect Tomato and Potato, Is Vectored by the Psyllid Bactericera cockerelli (Sulc) ▿
Article Snippet: Amplified DNA was cleaned using the Wizard PCR Preps DNA purification system (Promega, Madison, WI) and directly sequenced in both directions. .. Consequently, the four overlapping sets of primers designed by primer walking (see above and Table ) were used to amplify and subsequently sequence HLB bacteria DNA from plants.

Activation Assay:

Article Title: Differential effects of a common splice site polymorphism on the generation of OAS1 variants in human bronchial epithelial cells.
Article Snippet: The cycling conditions were as follows: initial activation of the Taq DNA Polymerase for 10 min at 95°C, followed by 40 cycles of 15 s denaturation at 95°C, and annealing and extension for 1 min at the appropriate temperature. .. Isoform-specific RT-PCR products containing the target sequences, amplified with primers in Supplementary Table 1, were purified using the Wizard PCR Preps DNA Purification System (Promega, Fitchburg, WI, USA).

Thin Layer Chromatography:

Article Title: Cloning and expression of 3-deoxy-d-manno-oct-2-ulosonic acid α-ketoside hydrolase from oyster hepatopancreas
Article Snippet: .. The following materials were purchased from commercial sources: marker proteins for molecular weight and p I , Sephacryl S-300 HR, Amersham Pharmacia Biotech; Tris, bis-Tris, MU-α-Neu5Ac and other MU-glycosides, Sigma; phenylmethylsulfonyl fluoride, Pierce; Silica Gel-60 thin-layer chromatography (TLC) plates, EMD Chemicals, Darmstadt, Germany; Centricon-10 and Microcon (10 kDa M r cut-off) micro-concentrators, Amicon; E. coli BL21(DE3), E. coli DH5α, S. cerevisiae INVSc1, P. pastoris , CHO-S cell, pUC18, pET, pYES2, pcDNA3.1/ myc -His(A), pPICZ, 5′-Race System for Rapid Amplification of cDNA Ends (Version 2) and LipofectAMINE 2000, Invitrogen; E. coli Origami, Novagen; GeneAmp PCR System 9600, Perkin-Elmer; Wizard PCR Preps DNA Purification System and pGEM-T Easy Vector System, Promega; RNAlater, Ambion; YPD and SD yeast culture media, BD Bioscience; FastTrack 2.0 Kit, BD Smart Race cDNA Amplification Kit and BD Advantage 2 PCR Kit, Clontech; Thermoscript RT-PCR System, Gibco; Oligotex Direct mRNA midi/maxi Kit and Ni-NTA Superflow column, Qiagen; chemically defined natural Re-LPS from S. minnesota and E. coli D31m4, Rd-LPS from S. minnesota R7 LIST Biological Laboratories, Inc; LPS from Neisseria meningitides strains A1 and M992 were gifts of Dr. Chao-Ming Tsai, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland. .. The α-Kdo-cleaving activity was measured by fluorometric analysis according to the procedure of ) as described previously ( ).

BAC Assay:

Article Title: A Deletion at the Mouse Xist Gene Exposes Trans-effects That Alter the Heterochromatin of the Inactive X Chromosome and the Replication Time and DNA Stability of Both X Chromosomes
Article Snippet: The mouse BAC RP23-412J16 [purchased from Invitrogen (Carlsbad, CA)] served as template for the PCR reactions. .. The PCR products were gel extracted using the Wizard PCR Preps DNA purification system (Promega).

Marker:

Article Title: Cloning and expression of 3-deoxy-d-manno-oct-2-ulosonic acid α-ketoside hydrolase from oyster hepatopancreas
Article Snippet: .. The following materials were purchased from commercial sources: marker proteins for molecular weight and p I , Sephacryl S-300 HR, Amersham Pharmacia Biotech; Tris, bis-Tris, MU-α-Neu5Ac and other MU-glycosides, Sigma; phenylmethylsulfonyl fluoride, Pierce; Silica Gel-60 thin-layer chromatography (TLC) plates, EMD Chemicals, Darmstadt, Germany; Centricon-10 and Microcon (10 kDa M r cut-off) micro-concentrators, Amicon; E. coli BL21(DE3), E. coli DH5α, S. cerevisiae INVSc1, P. pastoris , CHO-S cell, pUC18, pET, pYES2, pcDNA3.1/ myc -His(A), pPICZ, 5′-Race System for Rapid Amplification of cDNA Ends (Version 2) and LipofectAMINE 2000, Invitrogen; E. coli Origami, Novagen; GeneAmp PCR System 9600, Perkin-Elmer; Wizard PCR Preps DNA Purification System and pGEM-T Easy Vector System, Promega; RNAlater, Ambion; YPD and SD yeast culture media, BD Bioscience; FastTrack 2.0 Kit, BD Smart Race cDNA Amplification Kit and BD Advantage 2 PCR Kit, Clontech; Thermoscript RT-PCR System, Gibco; Oligotex Direct mRNA midi/maxi Kit and Ni-NTA Superflow column, Qiagen; chemically defined natural Re-LPS from S. minnesota and E. coli D31m4, Rd-LPS from S. minnesota R7 LIST Biological Laboratories, Inc; LPS from Neisseria meningitides strains A1 and M992 were gifts of Dr. Chao-Ming Tsai, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, Maryland. .. The α-Kdo-cleaving activity was measured by fluorometric analysis according to the procedure of ) as described previously ( ).

Fluorescence In Situ Hybridization:

Article Title: A Deletion at the Mouse Xist Gene Exposes Trans-effects That Alter the Heterochromatin of the Inactive X Chromosome and the Replication Time and DNA Stability of Both X Chromosomes
Article Snippet: Paragraph title: Two-color DNA FISH: ... The PCR products were gel extracted using the Wizard PCR Preps DNA purification system (Promega).

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    Promega wizard pcr preps dna purification system
    Restriction endonuclease digestion patterns for the A3′ amplicon of the CROP region of the toxin A gene for HSC98. The <t>PCR</t> amplicon product A3′ (lane 2) obtained from HSC98 using the A3 primer set (product was 1,250 bp) was exposed to Spe I (lane 3) and Eco I (lane 4) restriction endonucleases to determine restriction fragment length patterns. The A3′ amplicon (lane 2) had no restriction site for Eco RI (lane 4) and had one restriction site for Spe I (lane 3). A 500-bp <t>DNA</t> ladder is shown in lanes 1 and 6, and a 100-bp ladder is shown in lane 5.
    Wizard Pcr Preps Dna Purification System, supplied by Promega, used in various techniques. Bioz Stars score: 99/100, based on 205 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Restriction endonuclease digestion patterns for the A3′ amplicon of the CROP region of the toxin A gene for HSC98. The PCR amplicon product A3′ (lane 2) obtained from HSC98 using the A3 primer set (product was 1,250 bp) was exposed to Spe I (lane 3) and Eco I (lane 4) restriction endonucleases to determine restriction fragment length patterns. The A3′ amplicon (lane 2) had no restriction site for Eco RI (lane 4) and had one restriction site for Spe I (lane 3). A 500-bp DNA ladder is shown in lanes 1 and 6, and a 100-bp ladder is shown in lane 5.

    Journal: Journal of Clinical Microbiology

    Article Title: Characterization of a Toxin A-Negative, Toxin B-Positive Strain of Clostridium difficile Responsible for a Nosocomial Outbreak of Clostridium difficile-Associated Diarrhea

    doi:

    Figure Lengend Snippet: Restriction endonuclease digestion patterns for the A3′ amplicon of the CROP region of the toxin A gene for HSC98. The PCR amplicon product A3′ (lane 2) obtained from HSC98 using the A3 primer set (product was 1,250 bp) was exposed to Spe I (lane 3) and Eco I (lane 4) restriction endonucleases to determine restriction fragment length patterns. The A3′ amplicon (lane 2) had no restriction site for Eco RI (lane 4) and had one restriction site for Spe I (lane 3). A 500-bp DNA ladder is shown in lanes 1 and 6, and a 100-bp ladder is shown in lane 5.

    Article Snippet: The amplified product from this PCR protocol was purified using the Wizard PCR Preps DNA purification system (Promega Corp., Madison, Wis.), and this purified preparation was used for DNA sequencing.

    Techniques: Amplification, Polymerase Chain Reaction

    UP-PCR results of some patients. We have detected all the patients and finally we chose some positive bacteria and carried out the electrophoresis, so every line had PCR products. The deeper the stripe, the more the bacteria. Lines 1–9 represented PCR product of the 16S rRNA gene of Staphylococcus aureus , Staphylococcus epidermidis , Pseudomonas aeruginosa , Escherichia coli , Streptococcus viridans , Staphylococcus saprophyticus, Corynebacterium parvum , Klebsiella pneumoniae , and Enterobacter cloacae ; M for DL2000 DNA Marker; ten for negative control.

    Journal: BioMed Research International

    Article Title: Genetic Identification and Risk Factor Analysis of Asymptomatic Bacterial Colonization on Cardiovascular Implantable Electronic Devices

    doi: 10.1155/2014/725163

    Figure Lengend Snippet: UP-PCR results of some patients. We have detected all the patients and finally we chose some positive bacteria and carried out the electrophoresis, so every line had PCR products. The deeper the stripe, the more the bacteria. Lines 1–9 represented PCR product of the 16S rRNA gene of Staphylococcus aureus , Staphylococcus epidermidis , Pseudomonas aeruginosa , Escherichia coli , Streptococcus viridans , Staphylococcus saprophyticus, Corynebacterium parvum , Klebsiella pneumoniae , and Enterobacter cloacae ; M for DL2000 DNA Marker; ten for negative control.

    Article Snippet: The PCR product was purified using Wizard PCR Preps DNA Purification System (Promega) and then ligated into the pGEM-T Easy Vector (Promega).

    Techniques: Polymerase Chain Reaction, Electrophoresis, Marker, Negative Control

    The MALS mutagenesis procedure . Target DNA is individually amplified with two pairs of oligonucleotides SO1/IR1 and SO2/IF1. IF1 and IR1 are phosphorylated (rose circles), while SO1 and SO2 are dephosphorylated. The desired mutation is incorporated at the 5' end of IR1 oligonucleotide (shown in red). Panel 1 on the right shows the types of mutations available with MALS. PCR-generated DNA fragments are ligated. The resulting DNA population consists of homomeric ligation products (type A), non-ligated DNA fragments (type B), and molecules representing full length target DNA (type C) (panel 2 on the right). The entire DNA population is then amplified with suppression oligonucleotides SO1 and SO2. Intramolecular hybridization of inverted repeat sequences prevents efficient replication of type A molecules, while type B molecules amplify linearly (panel 3 on the right). Only heteromeric ligation products (type C) amplify exponentially. The resulting DNA population predominantly consists of type C molecules. An aliquot of the PCR mixture is used for the next round of mutagenesis employing internal oligonucleotides specific for another region of target DNA. Finally, SO1 and SO2 suppression sequences (green and blue, respectively) are digested with restriction endonucleases and DNA fragments are ligated with linearized vector for subsequent subcloning.

    Journal: BMC Biotechnology

    Article Title: MALS: an efficient strategy for multiple site-directed mutagenesis employing a combination of DNA amplification, ligation and suppression PCR

    doi: 10.1186/1472-6750-9-83

    Figure Lengend Snippet: The MALS mutagenesis procedure . Target DNA is individually amplified with two pairs of oligonucleotides SO1/IR1 and SO2/IF1. IF1 and IR1 are phosphorylated (rose circles), while SO1 and SO2 are dephosphorylated. The desired mutation is incorporated at the 5' end of IR1 oligonucleotide (shown in red). Panel 1 on the right shows the types of mutations available with MALS. PCR-generated DNA fragments are ligated. The resulting DNA population consists of homomeric ligation products (type A), non-ligated DNA fragments (type B), and molecules representing full length target DNA (type C) (panel 2 on the right). The entire DNA population is then amplified with suppression oligonucleotides SO1 and SO2. Intramolecular hybridization of inverted repeat sequences prevents efficient replication of type A molecules, while type B molecules amplify linearly (panel 3 on the right). Only heteromeric ligation products (type C) amplify exponentially. The resulting DNA population predominantly consists of type C molecules. An aliquot of the PCR mixture is used for the next round of mutagenesis employing internal oligonucleotides specific for another region of target DNA. Finally, SO1 and SO2 suppression sequences (green and blue, respectively) are digested with restriction endonucleases and DNA fragments are ligated with linearized vector for subsequent subcloning.

    Article Snippet: After amplification, the resulting DNA fragments were extracted from PCR mixtures using Wizard PCR Preps DNA Purification System (Promega, Madison, USA).

    Techniques: Mutagenesis, Amplification, Polymerase Chain Reaction, Generated, Ligation, Hybridization, Plasmid Preparation, Subcloning

    Agarose gel electrophoregram demonstrating the types of molecules forming during one individual round of mutagenesis . Lanes 1 and 2, PCR of target DNA with SO1/IR1 and SO2/IF1 oligonucleotides; Lane 3, homomeric (type A) and heteromeric (type C) DNA molecules formed during ligation of SO1/IR1 and SO2/IF1 DNA fragments (also, see Figure 1); Lane 4, suppression PCR with SO1 and SO2 oligonucleotides. The scheme on the right shows a graphical representation of type A and type C molecules that are present in lane 3 of the gel. During suppression PCR, intramolecular hybridization of inverted repeat sequences prevents binding of SO1 and SO2 oligonucleotides to the type A molecules that prevent their effective replication. Type C molecules amplify exponentially (see Additional File 1 for further details). Lane M, DNA size ladder with indicated positions of 1, 2, 3, 4, 6, and 10 kb DNA fragments (GeneRuler 1 kb DNA Ladder, Fermentas).

    Journal: BMC Biotechnology

    Article Title: MALS: an efficient strategy for multiple site-directed mutagenesis employing a combination of DNA amplification, ligation and suppression PCR

    doi: 10.1186/1472-6750-9-83

    Figure Lengend Snippet: Agarose gel electrophoregram demonstrating the types of molecules forming during one individual round of mutagenesis . Lanes 1 and 2, PCR of target DNA with SO1/IR1 and SO2/IF1 oligonucleotides; Lane 3, homomeric (type A) and heteromeric (type C) DNA molecules formed during ligation of SO1/IR1 and SO2/IF1 DNA fragments (also, see Figure 1); Lane 4, suppression PCR with SO1 and SO2 oligonucleotides. The scheme on the right shows a graphical representation of type A and type C molecules that are present in lane 3 of the gel. During suppression PCR, intramolecular hybridization of inverted repeat sequences prevents binding of SO1 and SO2 oligonucleotides to the type A molecules that prevent their effective replication. Type C molecules amplify exponentially (see Additional File 1 for further details). Lane M, DNA size ladder with indicated positions of 1, 2, 3, 4, 6, and 10 kb DNA fragments (GeneRuler 1 kb DNA Ladder, Fermentas).

    Article Snippet: After amplification, the resulting DNA fragments were extracted from PCR mixtures using Wizard PCR Preps DNA Purification System (Promega, Madison, USA).

    Techniques: Agarose Gel Electrophoresis, Mutagenesis, Polymerase Chain Reaction, Ligation, Hybridization, Binding Assay

    A. Agarose gel electrophoresis analysis of PCR product of HN of NDV, acmA, and HN-acmA Cassette ; Lane M: GeneRuler DNA ladder mix; Lanes 1: PCR product of HN of NDV, which is around ~ 1750 bp. Lanes 2: PCR product of acmA , which is around ~ 366 bp. Lanes 3: PCR product of HN-acmA , which is around ~ 2100 bp. B. SDS-PAGE and Western blot analysis of the purified recombinant HN-AcmA protein ; Lane N: Spectra Multicolor Broad Range Protein Ladder; Lane 1: pCDNA:HN-AcmA; Expression was confirmed through Western blot by using specific polyclonal antibody directed against NDV.

    Journal: Journal of Cancer

    Article Title: Newcastle Disease Virus Hemagglutinin Neuraminidase as a Potential Cancer Targeting Agent

    doi: 10.7150/jca.13566

    Figure Lengend Snippet: A. Agarose gel electrophoresis analysis of PCR product of HN of NDV, acmA, and HN-acmA Cassette ; Lane M: GeneRuler DNA ladder mix; Lanes 1: PCR product of HN of NDV, which is around ~ 1750 bp. Lanes 2: PCR product of acmA , which is around ~ 366 bp. Lanes 3: PCR product of HN-acmA , which is around ~ 2100 bp. B. SDS-PAGE and Western blot analysis of the purified recombinant HN-AcmA protein ; Lane N: Spectra Multicolor Broad Range Protein Ladder; Lane 1: pCDNA:HN-AcmA; Expression was confirmed through Western blot by using specific polyclonal antibody directed against NDV.

    Article Snippet: The ligation mixture was purified using the Wizard® PCR Preps DNA Purification Systems (Promega, USA) before the recombinant plasmids were transformed into FreeStyle™ CHO-S® cells.

    Techniques: Agarose Gel Electrophoresis, Polymerase Chain Reaction, SDS Page, Western Blot, Purification, Recombinant, Expressing