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Promega pgem t easy vector
Macrorestriction map of <t>pGEM-T</t> Easy-EGFRamiR. 1.1 kb Plus DNA Marker; 2. Restriction map of pGEM-T Easy-EGFRamiR by BamH I/BglII.
Pgem T Easy Vector, supplied by Promega, used in various techniques. Bioz Stars score: 99/100, based on 7378 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Images

1) Product Images from "The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR"

Article Title: The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR

Journal: International Journal of Clinical and Experimental Pathology

doi:

Macrorestriction map of pGEM-T Easy-EGFRamiR. 1.1 kb Plus DNA Marker; 2. Restriction map of pGEM-T Easy-EGFRamiR by BamH I/BglII.
Figure Legend Snippet: Macrorestriction map of pGEM-T Easy-EGFRamiR. 1.1 kb Plus DNA Marker; 2. Restriction map of pGEM-T Easy-EGFRamiR by BamH I/BglII.

Techniques Used: Marker

2) Product Images from "Screening and cloning for proteins transactivated by the PS1TP5 protein of hepatitis B virus: A suppression subtractive hybridization study"

Article Title: Screening and cloning for proteins transactivated by the PS1TP5 protein of hepatitis B virus: A suppression subtractive hybridization study

Journal:

doi: 10.3748/wjg.v13.i10.1602

Digestion and identification of the recombinant plasmid pGEM-T-PS1TP5 by Eco RI/ Bgl II. Lane 1: Restriction analysis of pGEM-T-PS1TP5; lane 2: DNA marker 2000.
Figure Legend Snippet: Digestion and identification of the recombinant plasmid pGEM-T-PS1TP5 by Eco RI/ Bgl II. Lane 1: Restriction analysis of pGEM-T-PS1TP5; lane 2: DNA marker 2000.

Techniques Used: Recombinant, Plasmid Preparation, Marker

3) Product Images from "Development and characterization of a novel CD34 monoclonal antibody that identifies sheep hematopoietic stem/progenitor cells"

Article Title: Development and characterization of a novel CD34 monoclonal antibody that identifies sheep hematopoietic stem/progenitor cells

Journal:

doi: 10.1016/j.exphem.2008.09.003

A) Nucleotide Sequence of the Extracellular Domain of Ovine CD34. The 858bp PCR product (see ) was excised from the gel, purified, and cloned into the pGEM–T Easy vector (Promega), as described in Materials and Methods. Sequencing was performed
Figure Legend Snippet: A) Nucleotide Sequence of the Extracellular Domain of Ovine CD34. The 858bp PCR product (see ) was excised from the gel, purified, and cloned into the pGEM–T Easy vector (Promega), as described in Materials and Methods. Sequencing was performed

Techniques Used: Sequencing, Polymerase Chain Reaction, Purification, Clone Assay, Plasmid Preparation

4) Product Images from "The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR"

Article Title: The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR

Journal: International Journal of Clinical and Experimental Pathology

doi:

Macrorestriction map of pGEM-T Easy-EGFRamiR. 1.1 kb Plus DNA Marker; 2. Restriction map of pGEM-T Easy-EGFRamiR by BamH I/BglII.
Figure Legend Snippet: Macrorestriction map of pGEM-T Easy-EGFRamiR. 1.1 kb Plus DNA Marker; 2. Restriction map of pGEM-T Easy-EGFRamiR by BamH I/BglII.

Techniques Used: Marker

5) Product Images from "The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR"

Article Title: The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR

Journal: International Journal of Clinical and Experimental Pathology

doi:

Macrorestriction map of pGEM-T Easy-EGFRamiR. 1.1 kb Plus DNA Marker; 2. Restriction map of pGEM-T Easy-EGFRamiR by BamH I/BglII.
Figure Legend Snippet: Macrorestriction map of pGEM-T Easy-EGFRamiR. 1.1 kb Plus DNA Marker; 2. Restriction map of pGEM-T Easy-EGFRamiR by BamH I/BglII.

Techniques Used: Marker

Related Articles

Transduction:

Article Title: The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR
Article Snippet: After 48 h or 72 h of transduction, the cell morphology was observed under a lighted microscope. .. The pDC312 vector, pBGHloxΔE1, 3Cre (Microbix Biosystems, Ontario, Canada), pGEM-T Easy vector (Promega, Beijing, China), adenovirus expression vector pDC312-SLPI-pA vector with SLPI promoter, and the BGH poly A sequence were constructed and saved by our lab.

Clone Assay:

Article Title: Characterization of White Spot Syndrome Virus Envelope Protein VP51A and Its Interaction with Viral Tegument Protein VP26 ▿
Article Snippet: .. The 5′ RACE products were cloned into the pGEM-T Easy vector, and seven clones were randomly chosen for sequencing with the results shown in Fig. . ..

Article Title: Two Arabidopsis ADP-Glucose Pyrophosphorylase Large Subunits (APL1 and APL2) Are Catalytic 1
Article Snippet: .. Absolute quantification ( ) was performed by cloning the amplified products in pGEM-T Easy vector (Promega) and using them as external calibration standards. .. Sequence data from this article can be found in the Arabidopsis Genome Initiative data library under accession numbers At5g48300 ( APS1 ), At1g05610 ( APS2 ), At5g19220 ( APL1 ), At1g27680 ( APL2 ), At4g39210 ( APL3 ), At2g21590 ( APL4 ), and SALK_040155 ( aps1 ).

Article Title: Role of proP and proU in Betaine Uptake by Yersinia enterocolitica under Cold and Osmotic Stress Conditions ▿ under Cold and Osmotic Stress Conditions ▿ †
Article Snippet: .. The amplified PCR products corresponding to the proP , proV , proW , and proX genes were gel extracted, cloned separately into the pGEM-T Easy vector (Promega Corp, Madison, WI) by utilizing the TA cloning strategy, and transformed into competent E. coli (JM109) cells (Promega) to obtain the recombinant plasmids pGEM proP , pGEM proV , pGEM proW , and pGEM proX . .. The presence of proP , proV , proW , and proX inserts in the recombinant white colonies was confirmed by restriction digestion and sequencing of the extracted plasmid DNA.

Article Title: Type II Thioesterase ScoT, Associated with Streptomyces coelicolor A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ †
Article Snippet: .. The PCR product was cloned in the pGEM-T Easy vector (Promega), digested with NdeI and HindIII, and cloned into the corresponding sites of pET-28a(+) to obtain plasmid pOS22. .. Mutated versions of the ScoT thioesterase were constructed by overlap extension ( ).

Article Title: The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR
Article Snippet: .. The EGFR amiR was cloned into a pGEM-T Easy vector for sequencing. .. After identification of the correct pGEM-T Easy-EGFR miRNA vector, the artificial EGFR miRNA with multiple repeat EGFR miRNA sequences was obtained by using BglII and BamHI restriction sites for the enhancement of the inhibitory effect of the target sequence.

Article Title: Screening and cloning for proteins transactivated by the PS1TP5 protein of hepatitis B virus: A suppression subtractive hybridization study
Article Snippet: Eukaryotic expression vector pcDNA3.1(-)-myc-his(A), Quickprep mico mRNA Purification kit, PCR-Select cDNA Subtraction kit, 50 × PCR Enzyme Mix kit and Advantage PCR Cloning kit were purchased from Clontech Co., America. .. FuGENE6 transfection kit was purchased from Roche Co., America. pGEM-T vector, pGEM-T-easy vector and High Pure PCR Product Purification kit were from Promega Co., America.

Article Title: The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR
Article Snippet: .. The EGFR amiR was cloned into the pGEM-T Easy vector. .. After identification of the correct pGEM-T Easy-EGFR miRNA vector, the artificial EGFR miRNA with multiple repeat EGFR miRNA sequences was obtained using BglII and BamHI restriction sites.

Article Title: NFAT but not NF-?B is critical for transcriptional induction of the prosurvival gene A1 after IgE receptor activation in mast cells
Article Snippet: .. The PCR product was cloned into the pGEM-T Easy vector (Promega, Madison, WI), confirmed by automated sequencing and finally subcloned into the pGL3-Basic vector (Promega) upstream of the luciferase reporter gene, generating the 1996pA1 Luc plasmid. .. The −389pA1 vector was constructed by deleting nucleotides −1996 to −390 from the −1996pA1 Luc plasmid.

Article Title: Development and characterization of a novel CD34 monoclonal antibody that identifies sheep hematopoietic stem/progenitor cells
Article Snippet: .. The 858bp band corresponding to the extracellular domain of the sheep CD34 gene was excised from the gel, purified using the Qiaex II kit (Qiagen, Inc., Valencia, CA) and cloned into the pGEM-T Easy vector (Promega, San Luis Obispo, CA), according to manufacturer's instructions. .. The recombinant plasmid was propagated in MAX Efficiency® DH5α™ Competent Cells (Invitrogen, Carslbad, CA) and purified using the QiaPrep Midiprep system (Qiagen).

Article Title: Transcription and genetic analyses of a putative N-acetylmuramyl-L-alanine amidase in Borrelia burgdorferi
Article Snippet: .. The obtained PCR product was then cloned into the pGEM-T Easy vector (Promega). .. Meanwhile, the full-length gfp gene was PCR amplified with engineered BamHI and PstI restriction at 5′ and 3′ ends, respectively.

Article Title: Satellite DNA From the Y Chromosome of the Malaria Vector Anopheles gambiae
Article Snippet: .. PCR reactions were performed in a Perkin-Elmer 9600 thermocycler with an initial denaturation at 94° for 3 min, followed by 35 cycles of 94° for 30 sec, 51°–55° for 30 sec, and 72° for 30–60 sec, followed by final elongation step at 72° for 10 min. PCR products were gel purified using a QIAquick Gel Extraction Kit (QIAGEN, Valencia, CA) and cloned into the pGEM-T Easy vector (Promega, Madison, WI). .. Cloned PCR templates were PCR amplified and gel purified prior to sequencing.

Amplification:

Article Title: Two Arabidopsis ADP-Glucose Pyrophosphorylase Large Subunits (APL1 and APL2) Are Catalytic 1
Article Snippet: .. Absolute quantification ( ) was performed by cloning the amplified products in pGEM-T Easy vector (Promega) and using them as external calibration standards. .. Sequence data from this article can be found in the Arabidopsis Genome Initiative data library under accession numbers At5g48300 ( APS1 ), At1g05610 ( APS2 ), At5g19220 ( APL1 ), At1g27680 ( APL2 ), At4g39210 ( APL3 ), At2g21590 ( APL4 ), and SALK_040155 ( aps1 ).

Article Title: Role of proP and proU in Betaine Uptake by Yersinia enterocolitica under Cold and Osmotic Stress Conditions ▿ under Cold and Osmotic Stress Conditions ▿ †
Article Snippet: .. The amplified PCR products corresponding to the proP , proV , proW , and proX genes were gel extracted, cloned separately into the pGEM-T Easy vector (Promega Corp, Madison, WI) by utilizing the TA cloning strategy, and transformed into competent E. coli (JM109) cells (Promega) to obtain the recombinant plasmids pGEM proP , pGEM proV , pGEM proW , and pGEM proX . .. The presence of proP , proV , proW , and proX inserts in the recombinant white colonies was confirmed by restriction digestion and sequencing of the extracted plasmid DNA.

Article Title: Type II Thioesterase ScoT, Associated with Streptomyces coelicolor A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ †
Article Snippet: The TE II gene scoT of S. coelicolor A3(2) was amplified by PCR using plasmid pMK15 as the template; pMK15 is a pBluescriptSK(+) construct containing two BamHI fragments from the T3 terminus of cosmid 1G7 (accession number ). .. The PCR product was cloned in the pGEM-T Easy vector (Promega), digested with NdeI and HindIII, and cloned into the corresponding sites of pET-28a(+) to obtain plasmid pOS22.

Article Title: NFAT but not NF-?B is critical for transcriptional induction of the prosurvival gene A1 after IgE receptor activation in mast cells
Article Snippet: The A1 upstream region encompassing nucleotides −1996 to +84 relative to the transcription start site was amplified by PCR from BALB/c genomic tail DNA using the following primers: 5′-GGGAGCGTGAAGAAGATATGG-3′, 5′-GCAAAGACTGCAAAGATGG-3′. .. The PCR product was cloned into the pGEM-T Easy vector (Promega, Madison, WI), confirmed by automated sequencing and finally subcloned into the pGL3-Basic vector (Promega) upstream of the luciferase reporter gene, generating the 1996pA1 Luc plasmid.

Article Title: Transcription and genetic analyses of a putative N-acetylmuramyl-L-alanine amidase in Borrelia burgdorferi
Article Snippet: To fuse Pami to gfp , the fragment containing Pami was PCR amplified using primers P24/P25 with engineered PstI and BamHI restriction cut sites at the 5′ and 3′ ends, respectively. .. The obtained PCR product was then cloned into the pGEM-T Easy vector (Promega).

Article Title: Satellite DNA From the Y Chromosome of the Malaria Vector Anopheles gambiae
Article Snippet: PCR reactions were performed in a Perkin-Elmer 9600 thermocycler with an initial denaturation at 94° for 3 min, followed by 35 cycles of 94° for 30 sec, 51°–55° for 30 sec, and 72° for 30–60 sec, followed by final elongation step at 72° for 10 min. PCR products were gel purified using a QIAquick Gel Extraction Kit (QIAGEN, Valencia, CA) and cloned into the pGEM-T Easy vector (Promega, Madison, WI). .. Cloned PCR templates were PCR amplified and gel purified prior to sequencing.

Synthesized:

Article Title: Role of proP and proU in Betaine Uptake by Yersinia enterocolitica under Cold and Osmotic Stress Conditions ▿ under Cold and Osmotic Stress Conditions ▿ †
Article Snippet: The primers for proP and individual ORFs of the proU operon were selected from the genome sequence of Y. pestis (GenBank accession no. ) and synthesized commercially (IDT DNA, Coralville, IA). .. The amplified PCR products corresponding to the proP , proV , proW , and proX genes were gel extracted, cloned separately into the pGEM-T Easy vector (Promega Corp, Madison, WI) by utilizing the TA cloning strategy, and transformed into competent E. coli (JM109) cells (Promega) to obtain the recombinant plasmids pGEM proP , pGEM proV , pGEM proW , and pGEM proX .

TA Cloning:

Article Title: Role of proP and proU in Betaine Uptake by Yersinia enterocolitica under Cold and Osmotic Stress Conditions ▿ under Cold and Osmotic Stress Conditions ▿ †
Article Snippet: .. The amplified PCR products corresponding to the proP , proV , proW , and proX genes were gel extracted, cloned separately into the pGEM-T Easy vector (Promega Corp, Madison, WI) by utilizing the TA cloning strategy, and transformed into competent E. coli (JM109) cells (Promega) to obtain the recombinant plasmids pGEM proP , pGEM proV , pGEM proW , and pGEM proX . .. The presence of proP , proV , proW , and proX inserts in the recombinant white colonies was confirmed by restriction digestion and sequencing of the extracted plasmid DNA.

Construct:

Article Title: Type II Thioesterase ScoT, Associated with Streptomyces coelicolor A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ †
Article Snippet: The TE II gene scoT of S. coelicolor A3(2) was amplified by PCR using plasmid pMK15 as the template; pMK15 is a pBluescriptSK(+) construct containing two BamHI fragments from the T3 terminus of cosmid 1G7 (accession number ). .. The PCR product was cloned in the pGEM-T Easy vector (Promega), digested with NdeI and HindIII, and cloned into the corresponding sites of pET-28a(+) to obtain plasmid pOS22.

Article Title: NFAT but not NF-?B is critical for transcriptional induction of the prosurvival gene A1 after IgE receptor activation in mast cells
Article Snippet: Paragraph title: Construction of A1 promoter-luciferase reporter constructs and NFAT vectors ... The PCR product was cloned into the pGEM-T Easy vector (Promega, Madison, WI), confirmed by automated sequencing and finally subcloned into the pGL3-Basic vector (Promega) upstream of the luciferase reporter gene, generating the 1996pA1 Luc plasmid.

Article Title: The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR
Article Snippet: .. The pDC312 vector, pBGHloxΔE1, 3Cre (Microbix Biosystems, Ontario, Canada), pGEM-T Easy vector (Promega, Beijing, China), adenovirus expression vector pDC312-SLPI-pA vector with SLPI promoter, and the BGH poly A sequence were constructed and saved by our lab. .. Using miR-30 as the basic framework, EGFR miRNA was designed according to the EGFR gene sequence (http://codex.cshl.edu/scripts/newmain.pl).

Article Title: Transcription and genetic analyses of a putative N-acetylmuramyl-L-alanine amidase in Borrelia burgdorferi
Article Snippet: Paragraph title: Fluorescent reporter constructs ... The obtained PCR product was then cloned into the pGEM-T Easy vector (Promega).

Real-time Polymerase Chain Reaction:

Article Title: Two Arabidopsis ADP-Glucose Pyrophosphorylase Large Subunits (APL1 and APL2) Are Catalytic 1
Article Snippet: Paragraph title: Real-Time PCR Analysis ... Absolute quantification ( ) was performed by cloning the amplified products in pGEM-T Easy vector (Promega) and using them as external calibration standards.

Luciferase:

Article Title: NFAT but not NF-?B is critical for transcriptional induction of the prosurvival gene A1 after IgE receptor activation in mast cells
Article Snippet: .. The PCR product was cloned into the pGEM-T Easy vector (Promega, Madison, WI), confirmed by automated sequencing and finally subcloned into the pGL3-Basic vector (Promega) upstream of the luciferase reporter gene, generating the 1996pA1 Luc plasmid. .. The −389pA1 vector was constructed by deleting nucleotides −1996 to −390 from the −1996pA1 Luc plasmid.

Introduce:

Article Title: Type II Thioesterase ScoT, Associated with Streptomyces coelicolor A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ †
Article Snippet: Primers TE-P-Nde (5′ TTT TTT TTT CAT ATG GGA AGT GAC TGG TT 3′) and TE-K-Hind (5′ TTT TTT T AA GCT T GT CGT ACG TAC ACG GA 3′) were designed to introduce restriction sites (underlined). .. The PCR product was cloned in the pGEM-T Easy vector (Promega), digested with NdeI and HindIII, and cloned into the corresponding sites of pET-28a(+) to obtain plasmid pOS22.

Expressing:

Article Title: Two Arabidopsis ADP-Glucose Pyrophosphorylase Large Subunits (APL1 and APL2) Are Catalytic 1
Article Snippet: Arabidopsis UBIQUITIN10 ( ) was used as a housekeeping gene control in the expression analysis. .. Absolute quantification ( ) was performed by cloning the amplified products in pGEM-T Easy vector (Promega) and using them as external calibration standards.

Article Title: Type II Thioesterase ScoT, Associated with Streptomyces coelicolor A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ †
Article Snippet: Paragraph title: Construction of the TE II expression plasmids. ... The PCR product was cloned in the pGEM-T Easy vector (Promega), digested with NdeI and HindIII, and cloned into the corresponding sites of pET-28a(+) to obtain plasmid pOS22.

Article Title: Screening and cloning for proteins transactivated by the PS1TP5 protein of hepatitis B virus: A suppression subtractive hybridization study
Article Snippet: Eukaryotic expression vector pcDNA3.1(-)-myc-his(A), Quickprep mico mRNA Purification kit, PCR-Select cDNA Subtraction kit, 50 × PCR Enzyme Mix kit and Advantage PCR Cloning kit were purchased from Clontech Co., America. .. FuGENE6 transfection kit was purchased from Roche Co., America. pGEM-T vector, pGEM-T-easy vector and High Pure PCR Product Purification kit were from Promega Co., America.

Article Title: The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR
Article Snippet: .. The pDC312 vector, pBGHloxΔE1, 3Cre (Microbix Biosystems, Ontario, Canada), pGEM-T Easy vector (Promega, Beijing, China), adenovirus expression vector pDC312-SLPI-pA vector with SLPI promoter, and the BGH poly A sequence were constructed and saved by our lab. .. Using miR-30 as the basic framework, EGFR miRNA was designed according to the EGFR gene sequence (http://codex.cshl.edu/scripts/newmain.pl).

Transformation Assay:

Article Title: Role of proP and proU in Betaine Uptake by Yersinia enterocolitica under Cold and Osmotic Stress Conditions ▿ under Cold and Osmotic Stress Conditions ▿ †
Article Snippet: .. The amplified PCR products corresponding to the proP , proV , proW , and proX genes were gel extracted, cloned separately into the pGEM-T Easy vector (Promega Corp, Madison, WI) by utilizing the TA cloning strategy, and transformed into competent E. coli (JM109) cells (Promega) to obtain the recombinant plasmids pGEM proP , pGEM proV , pGEM proW , and pGEM proX . .. The presence of proP , proV , proW , and proX inserts in the recombinant white colonies was confirmed by restriction digestion and sequencing of the extracted plasmid DNA.

Infection:

Article Title: The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR
Article Snippet: Hep-2 cells and HuVEC were seeded into 6-well plates at a density of 5 × 105 cells/well and maintained in RPMI1640 containing 10% fetal bovine serum for 24 h. Hep-2 cells and HuVEC were transduced with Ad-SLPI-EGFRamiR or Ad-SLPI-GFP at a multiplicity of infection (MOI) of 50 plaque forming units (pfu)/cell. .. The pDC312 vector, pBGHloxΔE1, 3Cre (Microbix Biosystems, Ontario, Canada), pGEM-T Easy vector (Promega, Beijing, China), adenovirus expression vector pDC312-SLPI-pA vector with SLPI promoter, and the BGH poly A sequence were constructed and saved by our lab.

Transfection:

Article Title: Screening and cloning for proteins transactivated by the PS1TP5 protein of hepatitis B virus: A suppression subtractive hybridization study
Article Snippet: .. FuGENE6 transfection kit was purchased from Roche Co., America. pGEM-T vector, pGEM-T-easy vector and High Pure PCR Product Purification kit were from Promega Co., America. .. Eco RI, BgI II and DNA Marker were from Takara Company, Japan.

Article Title: NFAT but not NF-?B is critical for transcriptional induction of the prosurvival gene A1 after IgE receptor activation in mast cells
Article Snippet: The PCR product was cloned into the pGEM-T Easy vector (Promega, Madison, WI), confirmed by automated sequencing and finally subcloned into the pGL3-Basic vector (Promega) upstream of the luciferase reporter gene, generating the 1996pA1 Luc plasmid. .. Reporter plasmids were confirmed by sequencing and prepared for transfection using Plasmid Midi Kit (QIAGEN, Hilden, Germany).

Cell Culture:

Article Title: The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR
Article Snippet: Paragraph title: Cells, cell culture, and reagents ... The pDC312 vector, pBGHloxΔE1, 3Cre (Microbix Biosystems, Ontario, Canada), pGEM-T Easy vector (Promega, Beijing, China), adenovirus expression vector pDC312-SLPI-pA vector with SLPI promoter, and the BGH poly A sequence were constructed and saved by our lab.

Generated:

Article Title: Role of proP and proU in Betaine Uptake by Yersinia enterocolitica under Cold and Osmotic Stress Conditions ▿ under Cold and Osmotic Stress Conditions ▿ †
Article Snippet: The amplified PCR products corresponding to the proP , proV , proW , and proX genes were gel extracted, cloned separately into the pGEM-T Easy vector (Promega Corp, Madison, WI) by utilizing the TA cloning strategy, and transformed into competent E. coli (JM109) cells (Promega) to obtain the recombinant plasmids pGEM proP , pGEM proV , pGEM proW , and pGEM proX . .. The sequences generated were analyzed by using Sequencher 4.1.4 (Gene Codes Corporation, Ann Arbor, MI), BPROM , ClustalW (EMBnet), and Blastn (NCBI).

Article Title: Type II Thioesterase ScoT, Associated with Streptomyces coelicolor A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ †
Article Snippet: The PCR product was cloned in the pGEM-T Easy vector (Promega), digested with NdeI and HindIII, and cloned into the corresponding sites of pET-28a(+) to obtain plasmid pOS22. .. Upstream and downstream portions of the scoT gene were amplified using Pfu polymerase, which generated blunt ends with appropriate primer pairs that introduced mutations (Table ).

Polymerase Chain Reaction:

Article Title: Two Arabidopsis ADP-Glucose Pyrophosphorylase Large Subunits (APL1 and APL2) Are Catalytic 1
Article Snippet: The PCR reaction mixture contained (in a total volume of 25 μ L): 5 μ L of cDNA, 0.2 m m dNTPs, 0.125 μ L of Exiqon Universal Probe Library (Roche), 1.25 μ L of 50 m m MgCl2 , 2.5 μ L of Ecotaq buffer 10× [670 m m Tris-HCl, pH 8.8, 166 m m (NH4 )2 SO4 , and 0.1% Tween 20], 0.3 units of EcoTaq polymerase (Ecogen), and 0.2 μ m of each primer. .. Absolute quantification ( ) was performed by cloning the amplified products in pGEM-T Easy vector (Promega) and using them as external calibration standards.

Article Title: Role of proP and proU in Betaine Uptake by Yersinia enterocolitica under Cold and Osmotic Stress Conditions ▿ under Cold and Osmotic Stress Conditions ▿ †
Article Snippet: .. The amplified PCR products corresponding to the proP , proV , proW , and proX genes were gel extracted, cloned separately into the pGEM-T Easy vector (Promega Corp, Madison, WI) by utilizing the TA cloning strategy, and transformed into competent E. coli (JM109) cells (Promega) to obtain the recombinant plasmids pGEM proP , pGEM proV , pGEM proW , and pGEM proX . .. The presence of proP , proV , proW , and proX inserts in the recombinant white colonies was confirmed by restriction digestion and sequencing of the extracted plasmid DNA.

Article Title: Type II Thioesterase ScoT, Associated with Streptomyces coelicolor A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ †
Article Snippet: .. The PCR product was cloned in the pGEM-T Easy vector (Promega), digested with NdeI and HindIII, and cloned into the corresponding sites of pET-28a(+) to obtain plasmid pOS22. .. Mutated versions of the ScoT thioesterase were constructed by overlap extension ( ).

Article Title: Screening and cloning for proteins transactivated by the PS1TP5 protein of hepatitis B virus: A suppression subtractive hybridization study
Article Snippet: .. FuGENE6 transfection kit was purchased from Roche Co., America. pGEM-T vector, pGEM-T-easy vector and High Pure PCR Product Purification kit were from Promega Co., America. .. Eco RI, BgI II and DNA Marker were from Takara Company, Japan.

Article Title: NFAT but not NF-?B is critical for transcriptional induction of the prosurvival gene A1 after IgE receptor activation in mast cells
Article Snippet: .. The PCR product was cloned into the pGEM-T Easy vector (Promega, Madison, WI), confirmed by automated sequencing and finally subcloned into the pGL3-Basic vector (Promega) upstream of the luciferase reporter gene, generating the 1996pA1 Luc plasmid. .. The −389pA1 vector was constructed by deleting nucleotides −1996 to −390 from the −1996pA1 Luc plasmid.

Article Title: Transcription and genetic analyses of a putative N-acetylmuramyl-L-alanine amidase in Borrelia burgdorferi
Article Snippet: .. The obtained PCR product was then cloned into the pGEM-T Easy vector (Promega). .. Meanwhile, the full-length gfp gene was PCR amplified with engineered BamHI and PstI restriction at 5′ and 3′ ends, respectively.

Article Title: Satellite DNA From the Y Chromosome of the Malaria Vector Anopheles gambiae
Article Snippet: .. PCR reactions were performed in a Perkin-Elmer 9600 thermocycler with an initial denaturation at 94° for 3 min, followed by 35 cycles of 94° for 30 sec, 51°–55° for 30 sec, and 72° for 30–60 sec, followed by final elongation step at 72° for 10 min. PCR products were gel purified using a QIAquick Gel Extraction Kit (QIAGEN, Valencia, CA) and cloned into the pGEM-T Easy vector (Promega, Madison, WI). .. Cloned PCR templates were PCR amplified and gel purified prior to sequencing.

Recombinant:

Article Title: Role of proP and proU in Betaine Uptake by Yersinia enterocolitica under Cold and Osmotic Stress Conditions ▿ under Cold and Osmotic Stress Conditions ▿ †
Article Snippet: .. The amplified PCR products corresponding to the proP , proV , proW , and proX genes were gel extracted, cloned separately into the pGEM-T Easy vector (Promega Corp, Madison, WI) by utilizing the TA cloning strategy, and transformed into competent E. coli (JM109) cells (Promega) to obtain the recombinant plasmids pGEM proP , pGEM proV , pGEM proW , and pGEM proX . .. The presence of proP , proV , proW , and proX inserts in the recombinant white colonies was confirmed by restriction digestion and sequencing of the extracted plasmid DNA.

Article Title: Development and characterization of a novel CD34 monoclonal antibody that identifies sheep hematopoietic stem/progenitor cells
Article Snippet: The 858bp band corresponding to the extracellular domain of the sheep CD34 gene was excised from the gel, purified using the Qiaex II kit (Qiagen, Inc., Valencia, CA) and cloned into the pGEM-T Easy vector (Promega, San Luis Obispo, CA), according to manufacturer's instructions. .. The recombinant plasmid was propagated in MAX Efficiency® DH5α™ Competent Cells (Invitrogen, Carslbad, CA) and purified using the QiaPrep Midiprep system (Qiagen).

Mutagenesis:

Article Title: Transcription and genetic analyses of a putative N-acetylmuramyl-L-alanine amidase in Borrelia burgdorferi
Article Snippet: To test whether Pami was functional, a mutant gfp gene was used as a transcriptional reporter ( ). .. The obtained PCR product was then cloned into the pGEM-T Easy vector (Promega).

Isolation:

Article Title: Development and characterization of a novel CD34 monoclonal antibody that identifies sheep hematopoietic stem/progenitor cells
Article Snippet: To obtain a cDNA clone for the extracellular domain of the sheep CD34 molecule, RNA purified from freshly isolated sheep dermal fibroblasts was provided by Dr. Paul Simmons and used to synthesize cDNA using the SuperScript® II First-Strand Synthesis System (Invitrogen, Carlsbad, CA). .. The 858bp band corresponding to the extracellular domain of the sheep CD34 gene was excised from the gel, purified using the Qiaex II kit (Qiagen, Inc., Valencia, CA) and cloned into the pGEM-T Easy vector (Promega, San Luis Obispo, CA), according to manufacturer's instructions.

Size-exclusion Chromatography:

Article Title: Satellite DNA From the Y Chromosome of the Malaria Vector Anopheles gambiae
Article Snippet: .. PCR reactions were performed in a Perkin-Elmer 9600 thermocycler with an initial denaturation at 94° for 3 min, followed by 35 cycles of 94° for 30 sec, 51°–55° for 30 sec, and 72° for 30–60 sec, followed by final elongation step at 72° for 10 min. PCR products were gel purified using a QIAquick Gel Extraction Kit (QIAGEN, Valencia, CA) and cloned into the pGEM-T Easy vector (Promega, Madison, WI). .. Cloned PCR templates were PCR amplified and gel purified prior to sequencing.

Microscopy:

Article Title: The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR
Article Snippet: After 48 h or 72 h of transduction, the cell morphology was observed under a lighted microscope. .. The pDC312 vector, pBGHloxΔE1, 3Cre (Microbix Biosystems, Ontario, Canada), pGEM-T Easy vector (Promega, Beijing, China), adenovirus expression vector pDC312-SLPI-pA vector with SLPI promoter, and the BGH poly A sequence were constructed and saved by our lab.

Purification:

Article Title: Screening and cloning for proteins transactivated by the PS1TP5 protein of hepatitis B virus: A suppression subtractive hybridization study
Article Snippet: .. FuGENE6 transfection kit was purchased from Roche Co., America. pGEM-T vector, pGEM-T-easy vector and High Pure PCR Product Purification kit were from Promega Co., America. .. Eco RI, BgI II and DNA Marker were from Takara Company, Japan.

Article Title: Development and characterization of a novel CD34 monoclonal antibody that identifies sheep hematopoietic stem/progenitor cells
Article Snippet: .. The 858bp band corresponding to the extracellular domain of the sheep CD34 gene was excised from the gel, purified using the Qiaex II kit (Qiagen, Inc., Valencia, CA) and cloned into the pGEM-T Easy vector (Promega, San Luis Obispo, CA), according to manufacturer's instructions. .. The recombinant plasmid was propagated in MAX Efficiency® DH5α™ Competent Cells (Invitrogen, Carslbad, CA) and purified using the QiaPrep Midiprep system (Qiagen).

Article Title: Satellite DNA From the Y Chromosome of the Malaria Vector Anopheles gambiae
Article Snippet: .. PCR reactions were performed in a Perkin-Elmer 9600 thermocycler with an initial denaturation at 94° for 3 min, followed by 35 cycles of 94° for 30 sec, 51°–55° for 30 sec, and 72° for 30–60 sec, followed by final elongation step at 72° for 10 min. PCR products were gel purified using a QIAquick Gel Extraction Kit (QIAGEN, Valencia, CA) and cloned into the pGEM-T Easy vector (Promega, Madison, WI). .. Cloned PCR templates were PCR amplified and gel purified prior to sequencing.

Sequencing:

Article Title: Characterization of White Spot Syndrome Virus Envelope Protein VP51A and Its Interaction with Viral Tegument Protein VP26 ▿
Article Snippet: .. The 5′ RACE products were cloned into the pGEM-T Easy vector, and seven clones were randomly chosen for sequencing with the results shown in Fig. . ..

Article Title: Role of proP and proU in Betaine Uptake by Yersinia enterocolitica under Cold and Osmotic Stress Conditions ▿ under Cold and Osmotic Stress Conditions ▿ †
Article Snippet: Paragraph title: Cloning and sequencing of proP , proV , proW , and proX of Y. enterocolitica . ... The amplified PCR products corresponding to the proP , proV , proW , and proX genes were gel extracted, cloned separately into the pGEM-T Easy vector (Promega Corp, Madison, WI) by utilizing the TA cloning strategy, and transformed into competent E. coli (JM109) cells (Promega) to obtain the recombinant plasmids pGEM proP , pGEM proV , pGEM proW , and pGEM proX .

Article Title: The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR
Article Snippet: .. The EGFR amiR was cloned into a pGEM-T Easy vector for sequencing. .. After identification of the correct pGEM-T Easy-EGFR miRNA vector, the artificial EGFR miRNA with multiple repeat EGFR miRNA sequences was obtained by using BglII and BamHI restriction sites for the enhancement of the inhibitory effect of the target sequence.

Article Title: NFAT but not NF-?B is critical for transcriptional induction of the prosurvival gene A1 after IgE receptor activation in mast cells
Article Snippet: .. The PCR product was cloned into the pGEM-T Easy vector (Promega, Madison, WI), confirmed by automated sequencing and finally subcloned into the pGL3-Basic vector (Promega) upstream of the luciferase reporter gene, generating the 1996pA1 Luc plasmid. .. The −389pA1 vector was constructed by deleting nucleotides −1996 to −390 from the −1996pA1 Luc plasmid.

Article Title: The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR
Article Snippet: .. The pDC312 vector, pBGHloxΔE1, 3Cre (Microbix Biosystems, Ontario, Canada), pGEM-T Easy vector (Promega, Beijing, China), adenovirus expression vector pDC312-SLPI-pA vector with SLPI promoter, and the BGH poly A sequence were constructed and saved by our lab. .. Using miR-30 as the basic framework, EGFR miRNA was designed according to the EGFR gene sequence (http://codex.cshl.edu/scripts/newmain.pl).

Article Title: Development and characterization of a novel CD34 monoclonal antibody that identifies sheep hematopoietic stem/progenitor cells
Article Snippet: Since the sheep CD34 gene had not been sequenced, these primers were designed based on the NCBI sequence for bovine CD34. .. The 858bp band corresponding to the extracellular domain of the sheep CD34 gene was excised from the gel, purified using the Qiaex II kit (Qiagen, Inc., Valencia, CA) and cloned into the pGEM-T Easy vector (Promega, San Luis Obispo, CA), according to manufacturer's instructions.

Article Title: Satellite DNA From the Y Chromosome of the Malaria Vector Anopheles gambiae
Article Snippet: Paragraph title: PCR, cloning, and sequencing: ... PCR reactions were performed in a Perkin-Elmer 9600 thermocycler with an initial denaturation at 94° for 3 min, followed by 35 cycles of 94° for 30 sec, 51°–55° for 30 sec, and 72° for 30–60 sec, followed by final elongation step at 72° for 10 min. PCR products were gel purified using a QIAquick Gel Extraction Kit (QIAGEN, Valencia, CA) and cloned into the pGEM-T Easy vector (Promega, Madison, WI).

IA:

Article Title: Role of proP and proU in Betaine Uptake by Yersinia enterocolitica under Cold and Osmotic Stress Conditions ▿ under Cold and Osmotic Stress Conditions ▿ †
Article Snippet: The primers for proP and individual ORFs of the proU operon were selected from the genome sequence of Y. pestis (GenBank accession no. ) and synthesized commercially (IDT DNA, Coralville, IA). .. The amplified PCR products corresponding to the proP , proV , proW , and proX genes were gel extracted, cloned separately into the pGEM-T Easy vector (Promega Corp, Madison, WI) by utilizing the TA cloning strategy, and transformed into competent E. coli (JM109) cells (Promega) to obtain the recombinant plasmids pGEM proP , pGEM proV , pGEM proW , and pGEM proX .

Chloramphenicol Acetyltransferase Assay:

Article Title: Type II Thioesterase ScoT, Associated with Streptomyces coelicolor A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ †
Article Snippet: Primers TE-P-Nde (5′ TTT TTT TTT CAT ATG GGA AGT GAC TGG TT 3′) and TE-K-Hind (5′ TTT TTT T AA GCT T GT CGT ACG TAC ACG GA 3′) were designed to introduce restriction sites (underlined). .. The PCR product was cloned in the pGEM-T Easy vector (Promega), digested with NdeI and HindIII, and cloned into the corresponding sites of pET-28a(+) to obtain plasmid pOS22.

Plasmid Preparation:

Article Title: Characterization of White Spot Syndrome Virus Envelope Protein VP51A and Its Interaction with Viral Tegument Protein VP26 ▿
Article Snippet: .. The 5′ RACE products were cloned into the pGEM-T Easy vector, and seven clones were randomly chosen for sequencing with the results shown in Fig. . ..

Article Title: Two Arabidopsis ADP-Glucose Pyrophosphorylase Large Subunits (APL1 and APL2) Are Catalytic 1
Article Snippet: .. Absolute quantification ( ) was performed by cloning the amplified products in pGEM-T Easy vector (Promega) and using them as external calibration standards. .. Sequence data from this article can be found in the Arabidopsis Genome Initiative data library under accession numbers At5g48300 ( APS1 ), At1g05610 ( APS2 ), At5g19220 ( APL1 ), At1g27680 ( APL2 ), At4g39210 ( APL3 ), At2g21590 ( APL4 ), and SALK_040155 ( aps1 ).

Article Title: Role of proP and proU in Betaine Uptake by Yersinia enterocolitica under Cold and Osmotic Stress Conditions ▿ under Cold and Osmotic Stress Conditions ▿ †
Article Snippet: .. The amplified PCR products corresponding to the proP , proV , proW , and proX genes were gel extracted, cloned separately into the pGEM-T Easy vector (Promega Corp, Madison, WI) by utilizing the TA cloning strategy, and transformed into competent E. coli (JM109) cells (Promega) to obtain the recombinant plasmids pGEM proP , pGEM proV , pGEM proW , and pGEM proX . .. The presence of proP , proV , proW , and proX inserts in the recombinant white colonies was confirmed by restriction digestion and sequencing of the extracted plasmid DNA.

Article Title: Type II Thioesterase ScoT, Associated with Streptomyces coelicolor A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ †
Article Snippet: .. The PCR product was cloned in the pGEM-T Easy vector (Promega), digested with NdeI and HindIII, and cloned into the corresponding sites of pET-28a(+) to obtain plasmid pOS22. .. Mutated versions of the ScoT thioesterase were constructed by overlap extension ( ).

Article Title: The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR
Article Snippet: .. The EGFR amiR was cloned into a pGEM-T Easy vector for sequencing. .. After identification of the correct pGEM-T Easy-EGFR miRNA vector, the artificial EGFR miRNA with multiple repeat EGFR miRNA sequences was obtained by using BglII and BamHI restriction sites for the enhancement of the inhibitory effect of the target sequence.

Article Title: Screening and cloning for proteins transactivated by the PS1TP5 protein of hepatitis B virus: A suppression subtractive hybridization study
Article Snippet: .. FuGENE6 transfection kit was purchased from Roche Co., America. pGEM-T vector, pGEM-T-easy vector and High Pure PCR Product Purification kit were from Promega Co., America. .. Eco RI, BgI II and DNA Marker were from Takara Company, Japan.

Article Title: The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR
Article Snippet: .. The EGFR amiR was cloned into the pGEM-T Easy vector. .. After identification of the correct pGEM-T Easy-EGFR miRNA vector, the artificial EGFR miRNA with multiple repeat EGFR miRNA sequences was obtained using BglII and BamHI restriction sites.

Article Title: NFAT but not NF-?B is critical for transcriptional induction of the prosurvival gene A1 after IgE receptor activation in mast cells
Article Snippet: .. The PCR product was cloned into the pGEM-T Easy vector (Promega, Madison, WI), confirmed by automated sequencing and finally subcloned into the pGL3-Basic vector (Promega) upstream of the luciferase reporter gene, generating the 1996pA1 Luc plasmid. .. The −389pA1 vector was constructed by deleting nucleotides −1996 to −390 from the −1996pA1 Luc plasmid.

Article Title: The construction and characterization of a novel adenovirus vector of artificial microRNA targeting EGFR
Article Snippet: .. The pDC312 vector, pBGHloxΔE1, 3Cre (Microbix Biosystems, Ontario, Canada), pGEM-T Easy vector (Promega, Beijing, China), adenovirus expression vector pDC312-SLPI-pA vector with SLPI promoter, and the BGH poly A sequence were constructed and saved by our lab. .. Using miR-30 as the basic framework, EGFR miRNA was designed according to the EGFR gene sequence (http://codex.cshl.edu/scripts/newmain.pl).

Article Title: Development and characterization of a novel CD34 monoclonal antibody that identifies sheep hematopoietic stem/progenitor cells
Article Snippet: .. The 858bp band corresponding to the extracellular domain of the sheep CD34 gene was excised from the gel, purified using the Qiaex II kit (Qiagen, Inc., Valencia, CA) and cloned into the pGEM-T Easy vector (Promega, San Luis Obispo, CA), according to manufacturer's instructions. .. The recombinant plasmid was propagated in MAX Efficiency® DH5α™ Competent Cells (Invitrogen, Carslbad, CA) and purified using the QiaPrep Midiprep system (Qiagen).

Article Title: Transcription and genetic analyses of a putative N-acetylmuramyl-L-alanine amidase in Borrelia burgdorferi
Article Snippet: .. The obtained PCR product was then cloned into the pGEM-T Easy vector (Promega). .. Meanwhile, the full-length gfp gene was PCR amplified with engineered BamHI and PstI restriction at 5′ and 3′ ends, respectively.

Article Title: Satellite DNA From the Y Chromosome of the Malaria Vector Anopheles gambiae
Article Snippet: .. PCR reactions were performed in a Perkin-Elmer 9600 thermocycler with an initial denaturation at 94° for 3 min, followed by 35 cycles of 94° for 30 sec, 51°–55° for 30 sec, and 72° for 30–60 sec, followed by final elongation step at 72° for 10 min. PCR products were gel purified using a QIAquick Gel Extraction Kit (QIAGEN, Valencia, CA) and cloned into the pGEM-T Easy vector (Promega, Madison, WI). .. Cloned PCR templates were PCR amplified and gel purified prior to sequencing.

Software:

Article Title: Satellite DNA From the Y Chromosome of the Malaria Vector Anopheles gambiae
Article Snippet: PCR reactions were performed in a Perkin-Elmer 9600 thermocycler with an initial denaturation at 94° for 3 min, followed by 35 cycles of 94° for 30 sec, 51°–55° for 30 sec, and 72° for 30–60 sec, followed by final elongation step at 72° for 10 min. PCR products were gel purified using a QIAquick Gel Extraction Kit (QIAGEN, Valencia, CA) and cloned into the pGEM-T Easy vector (Promega, Madison, WI). .. Sequences were assembled and verified by inspection of both strands using ABI Sequence Navigator software (Perkin-Elmer Applied Biosystems).

Functional Assay:

Article Title: Transcription and genetic analyses of a putative N-acetylmuramyl-L-alanine amidase in Borrelia burgdorferi
Article Snippet: To test whether Pami was functional, a mutant gfp gene was used as a transcriptional reporter ( ). .. The obtained PCR product was then cloned into the pGEM-T Easy vector (Promega).

Positron Emission Tomography:

Article Title: Type II Thioesterase ScoT, Associated with Streptomyces coelicolor A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ A3(2) Modular Polyketide Synthase Cpk, Hydrolyzes Acyl Residues and Has a Preference for Propionate ▿ †
Article Snippet: .. The PCR product was cloned in the pGEM-T Easy vector (Promega), digested with NdeI and HindIII, and cloned into the corresponding sites of pET-28a(+) to obtain plasmid pOS22. .. Mutated versions of the ScoT thioesterase were constructed by overlap extension ( ).

Marker:

Article Title: Screening and cloning for proteins transactivated by the PS1TP5 protein of hepatitis B virus: A suppression subtractive hybridization study
Article Snippet: FuGENE6 transfection kit was purchased from Roche Co., America. pGEM-T vector, pGEM-T-easy vector and High Pure PCR Product Purification kit were from Promega Co., America. .. Eco RI, BgI II and DNA Marker were from Takara Company, Japan.

Gel Extraction:

Article Title: Satellite DNA From the Y Chromosome of the Malaria Vector Anopheles gambiae
Article Snippet: .. PCR reactions were performed in a Perkin-Elmer 9600 thermocycler with an initial denaturation at 94° for 3 min, followed by 35 cycles of 94° for 30 sec, 51°–55° for 30 sec, and 72° for 30–60 sec, followed by final elongation step at 72° for 10 min. PCR products were gel purified using a QIAquick Gel Extraction Kit (QIAGEN, Valencia, CA) and cloned into the pGEM-T Easy vector (Promega, Madison, WI). .. Cloned PCR templates were PCR amplified and gel purified prior to sequencing.

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  • 99
    Promega pgem t easy cloning vector
    (A) Southern blotting of EcoRI-digested total DNA from FD-C-infected and FD-D-infected and healthy (H) periwinkles probed with a DIG-labeled malG gene amplicon obtained through <t>PCR</t> driven by primer pair malG _F/ malG _R (C+, probe positive control represented by <t>pGEM-T-</t> malG1 plasmid). (B) Electrophoresis separation of amplicons obtained following PCR of total DNA from FD-C-infected and FD-D-infected periwinkles with copy-specific primer pairs (002 and 005), according to the draft genome of FD92, and from healthy periwinkle. *, nonspecific PCR product.
    Pgem T Easy Cloning Vector, supplied by Promega, used in various techniques. Bioz Stars score: 99/100, based on 16 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pgem t easy cloning vector/product/Promega
    Average 99 stars, based on 16 article reviews
    Price from $9.99 to $1999.99
    pgem t easy cloning vector - by Bioz Stars, 2020-04
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    86
    Promega pax2 1 expression vector
    Expression of fadd is down-regulated in noi mutant zebrafish. ( A–F ) fadd expression in wild-type (wt), <t>pax2.1</t> −/− coloboma mutant ( noi ) and lamb1 −/− coloboma mutant ( gup ). Gene expression tested at either 24 (24
    Pax2 1 Expression Vector, supplied by Promega, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/pax2 1 expression vector/product/Promega
    Average 86 stars, based on 1 article reviews
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    pax2 1 expression vector - by Bioz Stars, 2020-04
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    86
    Promega β globin genes
    Fractionation of VSV-infected 293T cells. 293T cells were infected with VSV at different MOI (0.0001 to 0.1) and incubated for 16 hr. The nuclear and cytoplasmic fractions were separated and subjected to real-time PCR using the VSV-N (left panel) and <t>β-globin</t> primers (right panel). The DNA copy number per cell was estimated using the VSV N gene or β-globin gene in the control plasmid and is indicated on the axis. The means ± SE of two independent experiments performed in duplicate are shown.
    β Globin Genes, supplied by Promega, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Price from $9.99 to $1999.99
    β globin genes - by Bioz Stars, 2020-04
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    (A) Southern blotting of EcoRI-digested total DNA from FD-C-infected and FD-D-infected and healthy (H) periwinkles probed with a DIG-labeled malG gene amplicon obtained through PCR driven by primer pair malG _F/ malG _R (C+, probe positive control represented by pGEM-T- malG1 plasmid). (B) Electrophoresis separation of amplicons obtained following PCR of total DNA from FD-C-infected and FD-D-infected periwinkles with copy-specific primer pairs (002 and 005), according to the draft genome of FD92, and from healthy periwinkle. *, nonspecific PCR product.

    Journal: Applied and Environmental Microbiology

    Article Title: Genetic Diversity of Flavescence Dorée Phytoplasmas at the Vineyard Scale

    doi: 10.1128/AEM.03123-18

    Figure Lengend Snippet: (A) Southern blotting of EcoRI-digested total DNA from FD-C-infected and FD-D-infected and healthy (H) periwinkles probed with a DIG-labeled malG gene amplicon obtained through PCR driven by primer pair malG _F/ malG _R (C+, probe positive control represented by pGEM-T- malG1 plasmid). (B) Electrophoresis separation of amplicons obtained following PCR of total DNA from FD-C-infected and FD-D-infected periwinkles with copy-specific primer pairs (002 and 005), according to the draft genome of FD92, and from healthy periwinkle. *, nonspecific PCR product.

    Article Snippet: In the case of mixed infections (presence of double peaks in the analyzed pherograms from the sequencing of the original PCR amplicon), purified PCR products were ligated into pGEM-T easy cloning vector following the manufacturer's instructions (pGEM-T clone kit; Promega, Madison, WI) and transformed into Escherichia coli DH5α competent cells by heat shock.

    Techniques: Southern Blot, Infection, Labeling, Amplification, Polymerase Chain Reaction, Positive Control, Plasmid Preparation, Electrophoresis

    Amplification of the housekeeping genes and confirmation of cloning by EcoRI digestion of pGEM-T vector. (A) Amplification of the six target genes from cDNA of eggplant Lanes - M: 100 bp DNA Ladder/Marker, 1. 18S rRNA (416bp), 2: APRT (454 bp), 3: GAPDH (586bp), 4: Cyclophilin (265 bp), 5: Actin (333 bp), 6: RuBP (269bp). (B) Confirmation of the inserts in the recombinant pGEM-T vector by EcoRI digestion. Lanes - M: 100 bp DNA Ladder/Marker, 1. 18S rRNA (416bp), 2: APRT (454 bp), 3: Cyclophilin (265 bp), 4: GAPDH (586bp), 5: Actin (333 bp), 6: RuBP (269bp). (C) Amplification of the six target genes from genomic DNA of eggplant. Lanes- M: 100 bp DNA Ladder/Marker, 1. 18S rRNA (416bp), 2: APRT (No amplification), 3: Cyclophilin (265 bp), 4: GAPDH (1.2 kb), 5: Actin (333 bp), 6: RuBP (269bp).

    Journal: BMC Research Notes

    Article Title: Selection and validation of reference genes for quantitative gene expression studies by real-time PCR in eggplant (Solanum melongena L)

    doi: 10.1186/1756-0500-6-312

    Figure Lengend Snippet: Amplification of the housekeeping genes and confirmation of cloning by EcoRI digestion of pGEM-T vector. (A) Amplification of the six target genes from cDNA of eggplant Lanes - M: 100 bp DNA Ladder/Marker, 1. 18S rRNA (416bp), 2: APRT (454 bp), 3: GAPDH (586bp), 4: Cyclophilin (265 bp), 5: Actin (333 bp), 6: RuBP (269bp). (B) Confirmation of the inserts in the recombinant pGEM-T vector by EcoRI digestion. Lanes - M: 100 bp DNA Ladder/Marker, 1. 18S rRNA (416bp), 2: APRT (454 bp), 3: Cyclophilin (265 bp), 4: GAPDH (586bp), 5: Actin (333 bp), 6: RuBP (269bp). (C) Amplification of the six target genes from genomic DNA of eggplant. Lanes- M: 100 bp DNA Ladder/Marker, 1. 18S rRNA (416bp), 2: APRT (No amplification), 3: Cyclophilin (265 bp), 4: GAPDH (1.2 kb), 5: Actin (333 bp), 6: RuBP (269bp).

    Article Snippet: The amplified cDNA PCR products were cloned into pGEM-T Easy cloning vector (Promega) according to the manufacturer’s instructions.

    Techniques: Amplification, Clone Assay, Plasmid Preparation, Marker, Recombinant

    Expression of fadd is down-regulated in noi mutant zebrafish. ( A–F ) fadd expression in wild-type (wt), pax2.1 −/− coloboma mutant ( noi ) and lamb1 −/− coloboma mutant ( gup ). Gene expression tested at either 24 (24

    Journal: Human Molecular Genetics

    Article Title: Pax2 regulates a fadd-dependent molecular switch that drives tissue fusion during eye development

    doi: 10.1093/hmg/dds056

    Figure Lengend Snippet: Expression of fadd is down-regulated in noi mutant zebrafish. ( A–F ) fadd expression in wild-type (wt), pax2.1 −/− coloboma mutant ( noi ) and lamb1 −/− coloboma mutant ( gup ). Gene expression tested at either 24 (24

    Article Snippet: Full-length zebrafish pax2.1 protein was synthesized by in vitro translation from a pax2.1 expression vector (pGEM-T Easy) using the TNT T7-coupled reticulocyte lysate system (Promega).

    Techniques: Expressing, Mutagenesis

    Rescue of the pax2.1 -deficient phenotype by mRNA injection. ( A – E ) Wild-type embryos (wt) were injected with pax2.1 mRNA. ( F – J ) Uninjected pax2.1 -deficient (mt) embryos ( K – O ) pax2.1 -deficient embryos injected with pax2.1 mRNA. (

    Journal: Human Molecular Genetics

    Article Title: Pax2 regulates a fadd-dependent molecular switch that drives tissue fusion during eye development

    doi: 10.1093/hmg/dds056

    Figure Lengend Snippet: Rescue of the pax2.1 -deficient phenotype by mRNA injection. ( A – E ) Wild-type embryos (wt) were injected with pax2.1 mRNA. ( F – J ) Uninjected pax2.1 -deficient (mt) embryos ( K – O ) pax2.1 -deficient embryos injected with pax2.1 mRNA. (

    Article Snippet: Full-length zebrafish pax2.1 protein was synthesized by in vitro translation from a pax2.1 expression vector (pGEM-T Easy) using the TNT T7-coupled reticulocyte lysate system (Promega).

    Techniques: Injection

    Pax2.1 binds to the fadd promoter. ( A ) EMSA using fadd oligonucleotide probe containing the second pax2 -binding site. Fadd probe bound to the pax2.1 protein (lane 2), which was displaced by unlabelled competitor (lane 3) and in the presence of pax2 antibody

    Journal: Human Molecular Genetics

    Article Title: Pax2 regulates a fadd-dependent molecular switch that drives tissue fusion during eye development

    doi: 10.1093/hmg/dds056

    Figure Lengend Snippet: Pax2.1 binds to the fadd promoter. ( A ) EMSA using fadd oligonucleotide probe containing the second pax2 -binding site. Fadd probe bound to the pax2.1 protein (lane 2), which was displaced by unlabelled competitor (lane 3) and in the presence of pax2 antibody

    Article Snippet: Full-length zebrafish pax2.1 protein was synthesized by in vitro translation from a pax2.1 expression vector (pGEM-T Easy) using the TNT T7-coupled reticulocyte lysate system (Promega).

    Techniques: Binding Assay

    pax2.1 responsive elements in the fadd promoter. ( A ) Diagram at top depicts symbols used for transcription factor-binding sites. p, pax2.1 site; v, vax2 site; rxr, rxr site. Hatched box, exon 1 of fadd . L, luciferase gene. Reporter constructs transfected

    Journal: Human Molecular Genetics

    Article Title: Pax2 regulates a fadd-dependent molecular switch that drives tissue fusion during eye development

    doi: 10.1093/hmg/dds056

    Figure Lengend Snippet: pax2.1 responsive elements in the fadd promoter. ( A ) Diagram at top depicts symbols used for transcription factor-binding sites. p, pax2.1 site; v, vax2 site; rxr, rxr site. Hatched box, exon 1 of fadd . L, luciferase gene. Reporter constructs transfected

    Article Snippet: Full-length zebrafish pax2.1 protein was synthesized by in vitro translation from a pax2.1 expression vector (pGEM-T Easy) using the TNT T7-coupled reticulocyte lysate system (Promega).

    Techniques: Binding Assay, Luciferase, Construct, Transfection

    Spatiotemporal RIP3 localization and rescue of eye phenotype using necrostatin-1. ( A – C ) RIP3 immunohistochemistry (red) in wild-type (wt) eyes counterstained with DAPI. ( D and E ) High-level RIP3 labelling in pax2.1 -deficient eyes (mt). White arrow

    Journal: Human Molecular Genetics

    Article Title: Pax2 regulates a fadd-dependent molecular switch that drives tissue fusion during eye development

    doi: 10.1093/hmg/dds056

    Figure Lengend Snippet: Spatiotemporal RIP3 localization and rescue of eye phenotype using necrostatin-1. ( A – C ) RIP3 immunohistochemistry (red) in wild-type (wt) eyes counterstained with DAPI. ( D and E ) High-level RIP3 labelling in pax2.1 -deficient eyes (mt). White arrow

    Article Snippet: Full-length zebrafish pax2.1 protein was synthesized by in vitro translation from a pax2.1 expression vector (pGEM-T Easy) using the TNT T7-coupled reticulocyte lysate system (Promega).

    Techniques: Immunohistochemistry

    Fractionation of VSV-infected 293T cells. 293T cells were infected with VSV at different MOI (0.0001 to 0.1) and incubated for 16 hr. The nuclear and cytoplasmic fractions were separated and subjected to real-time PCR using the VSV-N (left panel) and β-globin primers (right panel). The DNA copy number per cell was estimated using the VSV N gene or β-globin gene in the control plasmid and is indicated on the axis. The means ± SE of two independent experiments performed in duplicate are shown.

    Journal: Scientific Reports

    Article Title: Characterisation of cytoplasmic DNA complementary to non-retroviral RNA viruses in human cells

    doi: 10.1038/srep05074

    Figure Lengend Snippet: Fractionation of VSV-infected 293T cells. 293T cells were infected with VSV at different MOI (0.0001 to 0.1) and incubated for 16 hr. The nuclear and cytoplasmic fractions were separated and subjected to real-time PCR using the VSV-N (left panel) and β-globin primers (right panel). The DNA copy number per cell was estimated using the VSV N gene or β-globin gene in the control plasmid and is indicated on the axis. The means ± SE of two independent experiments performed in duplicate are shown.

    Article Snippet: The copy number of each product was estimated using standard curves that were obtained using serial dilutions of the plasmids containing the VSV N and β-globin genes (pGEM®-T Easy vector) (Promega).

    Techniques: Fractionation, Infection, Incubation, Real-time Polymerase Chain Reaction, Plasmid Preparation

    LINE-1 transduction of the NP-2 cell line and its effect on VSV DNA formation. (a) LINE-1 transduction of the NP-2 cell line. The LINE-1 ORF1p and β-actin levels in the 293T, NP-2, NP-2/LINE-1 (L1.2), and NP-2/LINE-1 (L1.2 ORF2mut) cells were examined via Western blot using the anti-ORF1p IgY antibody. The ORF1p protein was observed at a molecular weight of ~41 kDa. The membrane was re-probed with an anti-β-actin antibody. The size marker at 38 kDa is indicated. Note that cropped western blots are shown and that full-length images are presented in the supplementary information . (b) The effect of LINE-1 transduction on VSV DNA production. The cells described above were infected with VSV, and the cell lysates were prepared. Real-time PCR was performed with either the N478-F/N681-R or β-globin primer pair. In addition, the VSV progeny produced by these cells were titrated and the VSV N mRNA expression levels were compared among these cells.

    Journal: Scientific Reports

    Article Title: Characterisation of cytoplasmic DNA complementary to non-retroviral RNA viruses in human cells

    doi: 10.1038/srep05074

    Figure Lengend Snippet: LINE-1 transduction of the NP-2 cell line and its effect on VSV DNA formation. (a) LINE-1 transduction of the NP-2 cell line. The LINE-1 ORF1p and β-actin levels in the 293T, NP-2, NP-2/LINE-1 (L1.2), and NP-2/LINE-1 (L1.2 ORF2mut) cells were examined via Western blot using the anti-ORF1p IgY antibody. The ORF1p protein was observed at a molecular weight of ~41 kDa. The membrane was re-probed with an anti-β-actin antibody. The size marker at 38 kDa is indicated. Note that cropped western blots are shown and that full-length images are presented in the supplementary information . (b) The effect of LINE-1 transduction on VSV DNA production. The cells described above were infected with VSV, and the cell lysates were prepared. Real-time PCR was performed with either the N478-F/N681-R or β-globin primer pair. In addition, the VSV progeny produced by these cells were titrated and the VSV N mRNA expression levels were compared among these cells.

    Article Snippet: The copy number of each product was estimated using standard curves that were obtained using serial dilutions of the plasmids containing the VSV N and β-globin genes (pGEM®-T Easy vector) (Promega).

    Techniques: Transduction, Western Blot, Molecular Weight, Marker, Infection, Real-time Polymerase Chain Reaction, Produced, Expressing

    Cell-based differences in VSV DNA production. Various human cells were infected with VSV at an MOI of 0.1. The DNA was extracted 16 hr later and subjected to PCR amplification. Real-time PCR was performed using the N478-F/N681-R or β-globin primer pair. The copy numbers of the VSV N and β-globin genes in each cell line are shown in closed and open bars, respectively. The arrows indicate that the copy number per cell was below the detection limit. The positive cells are ordered according to the VSV N DNA level, followed by the VSV DNA-negative adherent cells. Subsequently, the VSV DNA-negative suspension cells are arranged in alphabetical order. The means ± SE of two independent experiments performed in duplicate are shown.

    Journal: Scientific Reports

    Article Title: Characterisation of cytoplasmic DNA complementary to non-retroviral RNA viruses in human cells

    doi: 10.1038/srep05074

    Figure Lengend Snippet: Cell-based differences in VSV DNA production. Various human cells were infected with VSV at an MOI of 0.1. The DNA was extracted 16 hr later and subjected to PCR amplification. Real-time PCR was performed using the N478-F/N681-R or β-globin primer pair. The copy numbers of the VSV N and β-globin genes in each cell line are shown in closed and open bars, respectively. The arrows indicate that the copy number per cell was below the detection limit. The positive cells are ordered according to the VSV N DNA level, followed by the VSV DNA-negative adherent cells. Subsequently, the VSV DNA-negative suspension cells are arranged in alphabetical order. The means ± SE of two independent experiments performed in duplicate are shown.

    Article Snippet: The copy number of each product was estimated using standard curves that were obtained using serial dilutions of the plasmids containing the VSV N and β-globin genes (pGEM®-T Easy vector) (Promega).

    Techniques: Infection, Polymerase Chain Reaction, Amplification, Real-time Polymerase Chain Reaction