Transduction:Article Title: Vpx mediated degradation of SAMHD1 has only a very limited effect on lentiviral transduction rate in ex vivo cultured HSPCs
Article Snippet: .. THP-1 cells transduced with lentiviral vectors encoding SAMHD1 shRNA were selected by culturing in a medium containing 0.8 μg/ml puromycin. dNs consisted of a mixture of dA (D8668), dC (D0776), dG (D0901) and dT (T1895; all from Sigma-Aldrich). dNTPs (R0192) were purchased from Fermentas (Glen Burnie, MD). .. Cytokine cocktail for HSPC transduction consisted of a mixture of stem cell factor (SCF) at 100 ng/ml, Fms-related tyrosine kinase 3 ligand (Flt3L) at 100 ng/ml, interleukin-3 (IL-3) at 60 ng/ml and thrombopoietin (TPO) at 10 ng/ml (Immunotools, Friesoythe, Germany).
Amplification:Article Title: rpoB Gene Analysis as a Novel Strategy for Identification of Spirochetes from the Genera Borrelia, Treponema, and Leptospira
Article Snippet: .. Amplification of rpoB fragments was carried out by using the LTB primers listed in Table (3.2 pmol each) added to a 50-μl PCR mix containing 10 mM dNTP, 1 U of Taq polymerase (Gibco BRL), and 2 μl of DNA extract. ..
Article Title: Reconstituting ParA/ParB-mediated transport of DNA cargo
Article Snippet: .. pBR322:: sopC plasmid (PCR template) PCR Primers for sopC DNA amplification: (Primers synthesized by IDT) 10 mM dNTPs (Life Technologies; Cat. # 18427–013) 100 mM MgSO4 (New England Biolabs, Cat. # M0254S) 2000 U/mL VENT Polymerase (New England Biolabs, Cat. # M0254S) SYBR Gold DNA Stain (Life Technologies, Cat. # S-11494) Illustra MicroSpin S-400HR columns (GE Healthcare, Cat # 27–5140-01) MyOne Streptavidin C1 Dynabeads (Life Technologies, Cat. # 65001) 20 U/μL PstI Restriction Enzyme (New England Biolabs, Cat. # R0140S) Phenol:CHCl3:Isoamyl Alcohol (UltraPure 25:24:1 v/v) Ethanol. .. PCR Thermocycler Tabletop centrifuge Tube rotator BioMag Separator (Advanced Magnetics Inc.) Nanodrop Spectrophotometer.
Synthesized:Article Title: Reconstituting ParA/ParB-mediated transport of DNA cargo
Article Snippet: .. pBR322:: sopC plasmid (PCR template) PCR Primers for sopC DNA amplification: (Primers synthesized by IDT) 10 mM dNTPs (Life Technologies; Cat. # 18427–013) 100 mM MgSO4 (New England Biolabs, Cat. # M0254S) 2000 U/mL VENT Polymerase (New England Biolabs, Cat. # M0254S) SYBR Gold DNA Stain (Life Technologies, Cat. # S-11494) Illustra MicroSpin S-400HR columns (GE Healthcare, Cat # 27–5140-01) MyOne Streptavidin C1 Dynabeads (Life Technologies, Cat. # 65001) 20 U/μL PstI Restriction Enzyme (New England Biolabs, Cat. # R0140S) Phenol:CHCl3:Isoamyl Alcohol (UltraPure 25:24:1 v/v) Ethanol. .. PCR Thermocycler Tabletop centrifuge Tube rotator BioMag Separator (Advanced Magnetics Inc.) Nanodrop Spectrophotometer.
Protease Inhibitor:Article Title: Whole-genome chromatin profiling from limited numbers of cells using nano-ChIP-seq
Article Snippet: .. 25530-031) Protease inhibitor cocktail (Roche, Cat.#: 11836153001): Phenol:Chloroform:Isoamyl Alchohol (Invitrogen, Cat. #:15593031) RNAse (Invitrogen, Cat. #:12091-021) Sequenase Enzyme Kit (Version 2.0, US Biochemical, Cat. #: 70775 dNTP mix (Invitrogen, Cat.#: 18427-013 ) BSA (10 mg/ml , NEB, #B9001S) DTT (0.1 M stock, Invitrogen, #:Y00147) Primer 1 (4 μM): 5'-GACATGTATCCGGATGTNNNNNNNNN-3' ExoSAP-IT (USB, Cat.#:78200) Phusion Polymerase (NEB, Cat#: F-530L, provided with 100 % DMSO, 5X GC buffer) Primer 2 (10 μM stock): 5'-GACATGTATCCGGATGT-3’ BciVI Restriction Enzyme (NEB, Cat. #: R0596L) Agarose (Fisher, Cat #: BP160-500) Protein A-sepharose beads (Sigma, Cat#: P9424) PCR purification, Gel extraction and reaction clean up kits (Qiagen, Cat#s: 28006, 28606, 28206) Glycogen (20μg/μl, Invitrogen, Cat#: 10814-010) Quant-It dsDNA HS assay kit (Invitrogen, Cat#: ) .. 0.2 ml PCR tube strips (Eppendorf, Cat. #: 951010022) 1.5 ml tubes low retention tubes (Eppendorf, Cat. #:022431021) 15 ml tubes (BD Falcon cat. no. 352097) Filtered micropipette tips (Danville Sci., Cat.#s:P1096FR, P1121, P1122, P1126 ) Micropipiettes (1-1000 μl range) Spin-X wash columns (Costar, Cat#:8160) Refrigerated microcentrifuge (Eppendorf model 5415R or equivalent) Minicentrifuge with strip tubes adapter (Fisher Scientific Cat. #:05090-100) Cold room Sonicator (Branson 250 Digitial sonifier) Rotating mixer for 1.5 ml tubes Vortex mixer Thermal Cycler (Eppendorf, master cycle gradient or equivalent) Agarose gel electrophoresis equipment Qubit Flourometer (Invitrogen, Cat #: 32857)
Purification:Article Title: Whole-genome chromatin profiling from limited numbers of cells using nano-ChIP-seq
Article Snippet: .. 25530-031) Protease inhibitor cocktail (Roche, Cat.#: 11836153001): Phenol:Chloroform:Isoamyl Alchohol (Invitrogen, Cat. #:15593031) RNAse (Invitrogen, Cat. #:12091-021) Sequenase Enzyme Kit (Version 2.0, US Biochemical, Cat. #: 70775 dNTP mix (Invitrogen, Cat.#: 18427-013 ) BSA (10 mg/ml , NEB, #B9001S) DTT (0.1 M stock, Invitrogen, #:Y00147) Primer 1 (4 μM): 5'-GACATGTATCCGGATGTNNNNNNNNN-3' ExoSAP-IT (USB, Cat.#:78200) Phusion Polymerase (NEB, Cat#: F-530L, provided with 100 % DMSO, 5X GC buffer) Primer 2 (10 μM stock): 5'-GACATGTATCCGGATGT-3’ BciVI Restriction Enzyme (NEB, Cat. #: R0596L) Agarose (Fisher, Cat #: BP160-500) Protein A-sepharose beads (Sigma, Cat#: P9424) PCR purification, Gel extraction and reaction clean up kits (Qiagen, Cat#s: 28006, 28606, 28206) Glycogen (20μg/μl, Invitrogen, Cat#: 10814-010) Quant-It dsDNA HS assay kit (Invitrogen, Cat#: ) .. 0.2 ml PCR tube strips (Eppendorf, Cat. #: 951010022) 1.5 ml tubes low retention tubes (Eppendorf, Cat. #:022431021) 15 ml tubes (BD Falcon cat. no. 352097) Filtered micropipette tips (Danville Sci., Cat.#s:P1096FR, P1121, P1122, P1126 ) Micropipiettes (1-1000 μl range) Spin-X wash columns (Costar, Cat#:8160) Refrigerated microcentrifuge (Eppendorf model 5415R or equivalent) Minicentrifuge with strip tubes adapter (Fisher Scientific Cat. #:05090-100) Cold room Sonicator (Branson 250 Digitial sonifier) Rotating mixer for 1.5 ml tubes Vortex mixer Thermal Cycler (Eppendorf, master cycle gradient or equivalent) Agarose gel electrophoresis equipment Qubit Flourometer (Invitrogen, Cat #: 32857)
shRNA:Article Title: Vpx mediated degradation of SAMHD1 has only a very limited effect on lentiviral transduction rate in ex vivo cultured HSPCs
Article Snippet: .. THP-1 cells transduced with lentiviral vectors encoding SAMHD1 shRNA were selected by culturing in a medium containing 0.8 μg/ml puromycin. dNs consisted of a mixture of dA (D8668), dC (D0776), dG (D0901) and dT (T1895; all from Sigma-Aldrich). dNTPs (R0192) were purchased from Fermentas (Glen Burnie, MD). .. Cytokine cocktail for HSPC transduction consisted of a mixture of stem cell factor (SCF) at 100 ng/ml, Fms-related tyrosine kinase 3 ligand (Flt3L) at 100 ng/ml, interleukin-3 (IL-3) at 60 ng/ml and thrombopoietin (TPO) at 10 ng/ml (Immunotools, Friesoythe, Germany).
Lysis:Article Title: Direct Cell Lysis for Single-Cell Gene Expression Profiling
Article Snippet: .. The following chemicals were evaluated (final lysis concentrations are shown): 7-deaza-2′-deoxyguanosine-5′-triphosphate lithium salt (100 μM, Sigma-Aldrich); Betaine solution (4 M, Sigma-Aldrich); BSA (1–4 mg/ml, Fermentas); guanidine thiocyanate solution (GTC) (40–80 mM, Sigma-Aldrich); GenElute linear polyacrylamide (LPA) (50 ng/μl, Sigma-Aldrich); Igepal CA-630 (also known as Non-idet P-40, 0.5–4%, Sigma-Aldrich); polyinosinic acid potassium salt (50 ng/μl, Sigma-Aldrich); RNAseOUT (10 U/μl, Invitrogen); 2× reverse transcription buffer: 100 mM Tris-HCl (pH 8.3), 150 mM KCl, and 6 mM MgCl2 (Invitrogen); d -(+)-trehalose dihydrate (1 M, Sigma-Aldrich); yeast tRNA (50 ng/μl, Ambion); RT mix (2× RT buffer, Invitrogen, 5 μM random hexamers (Metabion), 5 μM oligo-dT (Metabion), 1 mM dNTP); RT mix with BSA (2× RT buffer, 5 μM random hexamers, 5 μM oligo-dT, 1 mM dNTP, 1 mg/ml BSA); and RNase-free water (Gibco). ..
Polymerase Chain Reaction:Article Title: A mutant Pfu DNA polymerase designed for advanced uracil-excision DNA engineering
Article Snippet: .. PCR with dUTP replacing dTTP was performed using standard Phusion (Finnzymes) reaction conditions as recommended by the manufacturer, using the dUTP/dNTPs mix (Fermentas) instead of dNTPs. .. The pET-Pfu-vector was a kind gift from Jens Lykke-Andersen.
Article Title: Whole-genome chromatin profiling from limited numbers of cells using nano-ChIP-seq
Article Snippet: .. 25530-031) Protease inhibitor cocktail (Roche, Cat.#: 11836153001): Phenol:Chloroform:Isoamyl Alchohol (Invitrogen, Cat. #:15593031) RNAse (Invitrogen, Cat. #:12091-021) Sequenase Enzyme Kit (Version 2.0, US Biochemical, Cat. #: 70775 dNTP mix (Invitrogen, Cat.#: 18427-013 ) BSA (10 mg/ml , NEB, #B9001S) DTT (0.1 M stock, Invitrogen, #:Y00147) Primer 1 (4 μM): 5'-GACATGTATCCGGATGTNNNNNNNNN-3' ExoSAP-IT (USB, Cat.#:78200) Phusion Polymerase (NEB, Cat#: F-530L, provided with 100 % DMSO, 5X GC buffer) Primer 2 (10 μM stock): 5'-GACATGTATCCGGATGT-3’ BciVI Restriction Enzyme (NEB, Cat. #: R0596L) Agarose (Fisher, Cat #: BP160-500) Protein A-sepharose beads (Sigma, Cat#: P9424) PCR purification, Gel extraction and reaction clean up kits (Qiagen, Cat#s: 28006, 28606, 28206) Glycogen (20μg/μl, Invitrogen, Cat#: 10814-010) Quant-It dsDNA HS assay kit (Invitrogen, Cat#: ) .. 0.2 ml PCR tube strips (Eppendorf, Cat. #: 951010022) 1.5 ml tubes low retention tubes (Eppendorf, Cat. #:022431021) 15 ml tubes (BD Falcon cat. no. 352097) Filtered micropipette tips (Danville Sci., Cat.#s:P1096FR, P1121, P1122, P1126 ) Micropipiettes (1-1000 μl range) Spin-X wash columns (Costar, Cat#:8160) Refrigerated microcentrifuge (Eppendorf model 5415R or equivalent) Minicentrifuge with strip tubes adapter (Fisher Scientific Cat. #:05090-100) Cold room Sonicator (Branson 250 Digitial sonifier) Rotating mixer for 1.5 ml tubes Vortex mixer Thermal Cycler (Eppendorf, master cycle gradient or equivalent) Agarose gel electrophoresis equipment Qubit Flourometer (Invitrogen, Cat #: 32857)
Article Title: Interleukin-17A is involved in mechanical hyperalgesia but not in the severity of murine antigen-induced arthritis
Article Snippet: .. The Master Mix consisted of 1x PCR buffer, 1 unit Taq DNA polymerase per 50 µl PCR mixture, 0.2 mM dNTP mixture (10 m M dNTP Mix; Thermo Fisher Scientific), and 0.5 µM primer. .. For amplification, 20–24 µl of Master Mix plus 1–5 µl cDNA template were used.
Article Title: rpoB Gene Analysis as a Novel Strategy for Identification of Spirochetes from the Genera Borrelia, Treponema, and Leptospira
Article Snippet: .. Amplification of rpoB fragments was carried out by using the LTB primers listed in Table (3.2 pmol each) added to a 50-μl PCR mix containing 10 mM dNTP, 1 U of Taq polymerase (Gibco BRL), and 2 μl of DNA extract. ..
Article Title: Reconstituting ParA/ParB-mediated transport of DNA cargo
Article Snippet: .. pBR322:: sopC plasmid (PCR template) PCR Primers for sopC DNA amplification: (Primers synthesized by IDT) 10 mM dNTPs (Life Technologies; Cat. # 18427–013) 100 mM MgSO4 (New England Biolabs, Cat. # M0254S) 2000 U/mL VENT Polymerase (New England Biolabs, Cat. # M0254S) SYBR Gold DNA Stain (Life Technologies, Cat. # S-11494) Illustra MicroSpin S-400HR columns (GE Healthcare, Cat # 27–5140-01) MyOne Streptavidin C1 Dynabeads (Life Technologies, Cat. # 65001) 20 U/μL PstI Restriction Enzyme (New England Biolabs, Cat. # R0140S) Phenol:CHCl3:Isoamyl Alcohol (UltraPure 25:24:1 v/v) Ethanol. .. PCR Thermocycler Tabletop centrifuge Tube rotator BioMag Separator (Advanced Magnetics Inc.) Nanodrop Spectrophotometer.
Article Title: Analysis of Endocannabinoid System in Rat Testis During the First Spermatogenetic Wave
Article Snippet: .. PCR was carried out using 2 µl cDNA and 10 pmol of the appropriate primers in a PCR mix [0.2 mM dNTP, 1× PCR buffer (Invitrogen Life Technologies), 1.5 mM MgCl2 , 1.25 U Taq Polymerase (Invitrogen Life Technologies, Paisley, UK)], using an Applied Biosystem Thermocycler. .. Possible residual genomic DNA was evaluated using Actin primers by RT− cDNA amplification (at 35 circles) while possible contamination among samples was excluded using samples prepared with 1 µl water (H2 O samples, negative control) in place of cDNA.
Gel Extraction:Article Title: Whole-genome chromatin profiling from limited numbers of cells using nano-ChIP-seq
Article Snippet: .. 25530-031) Protease inhibitor cocktail (Roche, Cat.#: 11836153001): Phenol:Chloroform:Isoamyl Alchohol (Invitrogen, Cat. #:15593031) RNAse (Invitrogen, Cat. #:12091-021) Sequenase Enzyme Kit (Version 2.0, US Biochemical, Cat. #: 70775 dNTP mix (Invitrogen, Cat.#: 18427-013 ) BSA (10 mg/ml , NEB, #B9001S) DTT (0.1 M stock, Invitrogen, #:Y00147) Primer 1 (4 μM): 5'-GACATGTATCCGGATGTNNNNNNNNN-3' ExoSAP-IT (USB, Cat.#:78200) Phusion Polymerase (NEB, Cat#: F-530L, provided with 100 % DMSO, 5X GC buffer) Primer 2 (10 μM stock): 5'-GACATGTATCCGGATGT-3’ BciVI Restriction Enzyme (NEB, Cat. #: R0596L) Agarose (Fisher, Cat #: BP160-500) Protein A-sepharose beads (Sigma, Cat#: P9424) PCR purification, Gel extraction and reaction clean up kits (Qiagen, Cat#s: 28006, 28606, 28206) Glycogen (20μg/μl, Invitrogen, Cat#: 10814-010) Quant-It dsDNA HS assay kit (Invitrogen, Cat#: ) .. 0.2 ml PCR tube strips (Eppendorf, Cat. #: 951010022) 1.5 ml tubes low retention tubes (Eppendorf, Cat. #:022431021) 15 ml tubes (BD Falcon cat. no. 352097) Filtered micropipette tips (Danville Sci., Cat.#s:P1096FR, P1121, P1122, P1126 ) Micropipiettes (1-1000 μl range) Spin-X wash columns (Costar, Cat#:8160) Refrigerated microcentrifuge (Eppendorf model 5415R or equivalent) Minicentrifuge with strip tubes adapter (Fisher Scientific Cat. #:05090-100) Cold room Sonicator (Branson 250 Digitial sonifier) Rotating mixer for 1.5 ml tubes Vortex mixer Thermal Cycler (Eppendorf, master cycle gradient or equivalent) Agarose gel electrophoresis equipment Qubit Flourometer (Invitrogen, Cat #: 32857)
Chloramphenicol Acetyltransferase Assay:Article Title: Whole-genome chromatin profiling from limited numbers of cells using nano-ChIP-seq
Article Snippet: .. 25530-031) Protease inhibitor cocktail (Roche, Cat.#: 11836153001): Phenol:Chloroform:Isoamyl Alchohol (Invitrogen, Cat. #:15593031) RNAse (Invitrogen, Cat. #:12091-021) Sequenase Enzyme Kit (Version 2.0, US Biochemical, Cat. #: 70775 dNTP mix (Invitrogen, Cat.#: 18427-013 ) BSA (10 mg/ml , NEB, #B9001S) DTT (0.1 M stock, Invitrogen, #:Y00147) Primer 1 (4 μM): 5'-GACATGTATCCGGATGTNNNNNNNNN-3' ExoSAP-IT (USB, Cat.#:78200) Phusion Polymerase (NEB, Cat#: F-530L, provided with 100 % DMSO, 5X GC buffer) Primer 2 (10 μM stock): 5'-GACATGTATCCGGATGT-3’ BciVI Restriction Enzyme (NEB, Cat. #: R0596L) Agarose (Fisher, Cat #: BP160-500) Protein A-sepharose beads (Sigma, Cat#: P9424) PCR purification, Gel extraction and reaction clean up kits (Qiagen, Cat#s: 28006, 28606, 28206) Glycogen (20μg/μl, Invitrogen, Cat#: 10814-010) Quant-It dsDNA HS assay kit (Invitrogen, Cat#: ) .. 0.2 ml PCR tube strips (Eppendorf, Cat. #: 951010022) 1.5 ml tubes low retention tubes (Eppendorf, Cat. #:022431021) 15 ml tubes (BD Falcon cat. no. 352097) Filtered micropipette tips (Danville Sci., Cat.#s:P1096FR, P1121, P1122, P1126 ) Micropipiettes (1-1000 μl range) Spin-X wash columns (Costar, Cat#:8160) Refrigerated microcentrifuge (Eppendorf model 5415R or equivalent) Minicentrifuge with strip tubes adapter (Fisher Scientific Cat. #:05090-100) Cold room Sonicator (Branson 250 Digitial sonifier) Rotating mixer for 1.5 ml tubes Vortex mixer Thermal Cycler (Eppendorf, master cycle gradient or equivalent) Agarose gel electrophoresis equipment Qubit Flourometer (Invitrogen, Cat #: 32857)
Plasmid Preparation:Article Title: Reconstituting ParA/ParB-mediated transport of DNA cargo
Article Snippet: .. pBR322:: sopC plasmid (PCR template) PCR Primers for sopC DNA amplification: (Primers synthesized by IDT) 10 mM dNTPs (Life Technologies; Cat. # 18427–013) 100 mM MgSO4 (New England Biolabs, Cat. # M0254S) 2000 U/mL VENT Polymerase (New England Biolabs, Cat. # M0254S) SYBR Gold DNA Stain (Life Technologies, Cat. # S-11494) Illustra MicroSpin S-400HR columns (GE Healthcare, Cat # 27–5140-01) MyOne Streptavidin C1 Dynabeads (Life Technologies, Cat. # 65001) 20 U/μL PstI Restriction Enzyme (New England Biolabs, Cat. # R0140S) Phenol:CHCl3:Isoamyl Alcohol (UltraPure 25:24:1 v/v) Ethanol. .. PCR Thermocycler Tabletop centrifuge Tube rotator BioMag Separator (Advanced Magnetics Inc.) Nanodrop Spectrophotometer.
Staining:Article Title: Reconstituting ParA/ParB-mediated transport of DNA cargo
Article Snippet: .. pBR322:: sopC plasmid (PCR template) PCR Primers for sopC DNA amplification: (Primers synthesized by IDT) 10 mM dNTPs (Life Technologies; Cat. # 18427–013) 100 mM MgSO4 (New England Biolabs, Cat. # M0254S) 2000 U/mL VENT Polymerase (New England Biolabs, Cat. # M0254S) SYBR Gold DNA Stain (Life Technologies, Cat. # S-11494) Illustra MicroSpin S-400HR columns (GE Healthcare, Cat # 27–5140-01) MyOne Streptavidin C1 Dynabeads (Life Technologies, Cat. # 65001) 20 U/μL PstI Restriction Enzyme (New England Biolabs, Cat. # R0140S) Phenol:CHCl3:Isoamyl Alcohol (UltraPure 25:24:1 v/v) Ethanol. .. PCR Thermocycler Tabletop centrifuge Tube rotator BioMag Separator (Advanced Magnetics Inc.) Nanodrop Spectrophotometer.
|
Thermo Fisher is a verified supplier 