Structured Review

ATUM input dna
LPS and TNFα induce nuclear localization and binding of Runx proteins to the CD40 promoter. (A) Schematic presentation of mouse CD40 promoter indicating the location of putative Runx-binding sites (R1 and R2) and ChIP primers (P1 and P2), and details of oligonucleotide probes used for EMSA. Mouse CD40 promoter-specific Pr1 and Pr2 probes contain putative wild-type Runx-binding sites, and Mut-Pr1 and Mut-Pr2 probes contain mutations (in italics) at Runx-binding sites. Base positions are relative to the transcription start site. (B and C) EMSA of nuclear extracts of BMDCs treated with LPS or TNFα for indicated times; assessed with indicated probes. Numbers below lanes in (B) represent densitometry [normalized to OCT-1 binding (control)] relative to that of untreated BMDCs (0 h). The densitometry results pooled from three independent experiments for (B) are shown in S2 Fig . (D) Supershift EMSA [with immunoglobulin G (IgG; control) or antibody to (α-) Runx1, Runx2 or Runx3] of nuclear extracts of BMDCs treated with LPS or TNFα for 0.5 h; assayed with Pr1 or Pr2 probes. (E) Binding of Runx proteins to -578/-373 region of the CD40 promoter in BMDCs treated with LPS or TNFα (time, above lanes); assessed via ChIP using the primers shown in (A) and indicated antibodies (left margin). Amplification of mouse GAPDH promoter ( S4 Fig ) and chromatin <t>immunoprecipitated</t> by rabbit IgG were used as negative controls, and input <t>DNA</t> (2%) as an internal control. Numbers below lanes represent densitometry, normalized to input DNA and presented relative to that of untreated BMDCs (0 h). (F) In vivo footprint analysis of mouse CD40 promoter showing DMS-protected bands (indicated by open circles) at R1 and R2 sites in LPS- or TNFα-treated but not in untreated (UT) BMDCs. (G) Translocation of Runx1 or Runx3 (red) into the nuclei (blue; Hoechst staining) in BMDCs left untreated (UT) or treated with LPS or TNFα for 0.5 h; assessed by confocal microscopy. Pink color (merge) shows nuclear translocation of Runx1 or Runx3. Scale bar, 10 μm. Confocal microscopy images of DCs immunostained with isotype control antibodies are given in S5 Fig . Data are representative of three (B-E) or two (F and G) independent experiments.
Input Dna, supplied by ATUM, 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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Images

1) Product Images from "Runx proteins mediate protective immunity against Leishmania donovani infection by promoting CD40 expression on dendritic cells"

Article Title: Runx proteins mediate protective immunity against Leishmania donovani infection by promoting CD40 expression on dendritic cells

Journal: PLoS Pathogens

doi: 10.1371/journal.ppat.1009136

LPS and TNFα induce nuclear localization and binding of Runx proteins to the CD40 promoter. (A) Schematic presentation of mouse CD40 promoter indicating the location of putative Runx-binding sites (R1 and R2) and ChIP primers (P1 and P2), and details of oligonucleotide probes used for EMSA. Mouse CD40 promoter-specific Pr1 and Pr2 probes contain putative wild-type Runx-binding sites, and Mut-Pr1 and Mut-Pr2 probes contain mutations (in italics) at Runx-binding sites. Base positions are relative to the transcription start site. (B and C) EMSA of nuclear extracts of BMDCs treated with LPS or TNFα for indicated times; assessed with indicated probes. Numbers below lanes in (B) represent densitometry [normalized to OCT-1 binding (control)] relative to that of untreated BMDCs (0 h). The densitometry results pooled from three independent experiments for (B) are shown in S2 Fig . (D) Supershift EMSA [with immunoglobulin G (IgG; control) or antibody to (α-) Runx1, Runx2 or Runx3] of nuclear extracts of BMDCs treated with LPS or TNFα for 0.5 h; assayed with Pr1 or Pr2 probes. (E) Binding of Runx proteins to -578/-373 region of the CD40 promoter in BMDCs treated with LPS or TNFα (time, above lanes); assessed via ChIP using the primers shown in (A) and indicated antibodies (left margin). Amplification of mouse GAPDH promoter ( S4 Fig ) and chromatin immunoprecipitated by rabbit IgG were used as negative controls, and input DNA (2%) as an internal control. Numbers below lanes represent densitometry, normalized to input DNA and presented relative to that of untreated BMDCs (0 h). (F) In vivo footprint analysis of mouse CD40 promoter showing DMS-protected bands (indicated by open circles) at R1 and R2 sites in LPS- or TNFα-treated but not in untreated (UT) BMDCs. (G) Translocation of Runx1 or Runx3 (red) into the nuclei (blue; Hoechst staining) in BMDCs left untreated (UT) or treated with LPS or TNFα for 0.5 h; assessed by confocal microscopy. Pink color (merge) shows nuclear translocation of Runx1 or Runx3. Scale bar, 10 μm. Confocal microscopy images of DCs immunostained with isotype control antibodies are given in S5 Fig . Data are representative of three (B-E) or two (F and G) independent experiments.
Figure Legend Snippet: LPS and TNFα induce nuclear localization and binding of Runx proteins to the CD40 promoter. (A) Schematic presentation of mouse CD40 promoter indicating the location of putative Runx-binding sites (R1 and R2) and ChIP primers (P1 and P2), and details of oligonucleotide probes used for EMSA. Mouse CD40 promoter-specific Pr1 and Pr2 probes contain putative wild-type Runx-binding sites, and Mut-Pr1 and Mut-Pr2 probes contain mutations (in italics) at Runx-binding sites. Base positions are relative to the transcription start site. (B and C) EMSA of nuclear extracts of BMDCs treated with LPS or TNFα for indicated times; assessed with indicated probes. Numbers below lanes in (B) represent densitometry [normalized to OCT-1 binding (control)] relative to that of untreated BMDCs (0 h). The densitometry results pooled from three independent experiments for (B) are shown in S2 Fig . (D) Supershift EMSA [with immunoglobulin G (IgG; control) or antibody to (α-) Runx1, Runx2 or Runx3] of nuclear extracts of BMDCs treated with LPS or TNFα for 0.5 h; assayed with Pr1 or Pr2 probes. (E) Binding of Runx proteins to -578/-373 region of the CD40 promoter in BMDCs treated with LPS or TNFα (time, above lanes); assessed via ChIP using the primers shown in (A) and indicated antibodies (left margin). Amplification of mouse GAPDH promoter ( S4 Fig ) and chromatin immunoprecipitated by rabbit IgG were used as negative controls, and input DNA (2%) as an internal control. Numbers below lanes represent densitometry, normalized to input DNA and presented relative to that of untreated BMDCs (0 h). (F) In vivo footprint analysis of mouse CD40 promoter showing DMS-protected bands (indicated by open circles) at R1 and R2 sites in LPS- or TNFα-treated but not in untreated (UT) BMDCs. (G) Translocation of Runx1 or Runx3 (red) into the nuclei (blue; Hoechst staining) in BMDCs left untreated (UT) or treated with LPS or TNFα for 0.5 h; assessed by confocal microscopy. Pink color (merge) shows nuclear translocation of Runx1 or Runx3. Scale bar, 10 μm. Confocal microscopy images of DCs immunostained with isotype control antibodies are given in S5 Fig . Data are representative of three (B-E) or two (F and G) independent experiments.

Techniques Used: Binding Assay, Chromatin Immunoprecipitation, Amplification, Immunoprecipitation, In Vivo, Translocation Assay, Staining, Confocal Microscopy

Sb R LD inhibits SAG-induced CD40 upregulation on HuMoDCs by impairing Runx binding to the CD40 promoter. (A) A schematic of human CD40 promoter showing the location of putative Runx-binding sites (R3 and R4) and ChIP primers (P3 and P4), and details of oligonucleotide probes used for EMSA. Human CD40 promoter-specific Pr3 and Pr4 probes contain putative wild-type Runx-binding sites, and Mut-Pr3 and Mut-Pr4 probes contain mutations (in italics) at Runx-binding sites. Base positions are relative to the transcription start site. (B and C) EMSA of nuclear extracts of HuMoDCs treated with SAG for indicated times; assayed with indicated probes. OCT-1 binding (in B) serves as an internal control. Numbers below lanes in (B), densitometry readings (as in Fig 2B ); presented relative to untreated HuMoDCs (0 h). (D) Supershift EMSA (antibodies and probes are indicated above and below lanes, respectively) to assess the binding of nuclear Runx1, Runx2 and Runx3 to the CD40 promoter in HuMoDCs treated with SAG for 0.5 h. (E) ChIP assay [with the primers shown in (A) and antibodies indicated at left margin] to determine the binding of Runx proteins to -480/-276 region of the CD40 promoter in HuMoDCs treated with SAG (time, above lanes). Amplification of human GAPDH promoter and chromatin immunoprecipitated by rabbit IgG were used as negative controls, and input DNA (2%) as an internal control. Numbers below lanes represent densitometry, normalized to input DNA and presented relative to that of untreated HuMoDCs (0 h). hu CD40 , human CD40 ; hu GAPDH , human GAPDH . (F) Immunoblot analysis of Runx1 and Runx3 expression in HuMoDCs left untransfected, or transfected with control siRNA, Runx1 siRNA, Runx3 siRNA or Runx1 and Runx3 siRNAs (Runx1+3 siRNA). β-actin serves as a loading control. (G) Flow cytometry analysis of CD40 expression on HuMoDCs transfected with indicated siRNAs and then cultured with or without SAG for 24 h. (H) EMSA of nuclear extracts of HuMoDCs left uninfected or infected with promastigotes of Sb R LD strain GE1F8R or Sb S LD strain AG83 for 24 h and then cultured for 0.5 h with (+) or without (-) SAG; assayed with indicated probes. OCT-1 binding serves as an internal control. Numbers below lanes represent densitometry readings (as in Fig 2B ); presented relative to control HuMoDCs (HuMoDCs left uninfected and cultured without SAG). (I) HuMoDCs were either left uninfected or infected with promastigotes of Sb S LD strain AG83 or Sb R LD strain GE1F8R for 24 h and then treated (for 24 h) with or without SAG. CD40 expression on HuMoDCs was assessed by flow cytometry. All data are representative of two independent experiments.
Figure Legend Snippet: Sb R LD inhibits SAG-induced CD40 upregulation on HuMoDCs by impairing Runx binding to the CD40 promoter. (A) A schematic of human CD40 promoter showing the location of putative Runx-binding sites (R3 and R4) and ChIP primers (P3 and P4), and details of oligonucleotide probes used for EMSA. Human CD40 promoter-specific Pr3 and Pr4 probes contain putative wild-type Runx-binding sites, and Mut-Pr3 and Mut-Pr4 probes contain mutations (in italics) at Runx-binding sites. Base positions are relative to the transcription start site. (B and C) EMSA of nuclear extracts of HuMoDCs treated with SAG for indicated times; assayed with indicated probes. OCT-1 binding (in B) serves as an internal control. Numbers below lanes in (B), densitometry readings (as in Fig 2B ); presented relative to untreated HuMoDCs (0 h). (D) Supershift EMSA (antibodies and probes are indicated above and below lanes, respectively) to assess the binding of nuclear Runx1, Runx2 and Runx3 to the CD40 promoter in HuMoDCs treated with SAG for 0.5 h. (E) ChIP assay [with the primers shown in (A) and antibodies indicated at left margin] to determine the binding of Runx proteins to -480/-276 region of the CD40 promoter in HuMoDCs treated with SAG (time, above lanes). Amplification of human GAPDH promoter and chromatin immunoprecipitated by rabbit IgG were used as negative controls, and input DNA (2%) as an internal control. Numbers below lanes represent densitometry, normalized to input DNA and presented relative to that of untreated HuMoDCs (0 h). hu CD40 , human CD40 ; hu GAPDH , human GAPDH . (F) Immunoblot analysis of Runx1 and Runx3 expression in HuMoDCs left untransfected, or transfected with control siRNA, Runx1 siRNA, Runx3 siRNA or Runx1 and Runx3 siRNAs (Runx1+3 siRNA). β-actin serves as a loading control. (G) Flow cytometry analysis of CD40 expression on HuMoDCs transfected with indicated siRNAs and then cultured with or without SAG for 24 h. (H) EMSA of nuclear extracts of HuMoDCs left uninfected or infected with promastigotes of Sb R LD strain GE1F8R or Sb S LD strain AG83 for 24 h and then cultured for 0.5 h with (+) or without (-) SAG; assayed with indicated probes. OCT-1 binding serves as an internal control. Numbers below lanes represent densitometry readings (as in Fig 2B ); presented relative to control HuMoDCs (HuMoDCs left uninfected and cultured without SAG). (I) HuMoDCs were either left uninfected or infected with promastigotes of Sb S LD strain AG83 or Sb R LD strain GE1F8R for 24 h and then treated (for 24 h) with or without SAG. CD40 expression on HuMoDCs was assessed by flow cytometry. All data are representative of two independent experiments.

Techniques Used: Binding Assay, Chromatin Immunoprecipitation, Amplification, Immunoprecipitation, Expressing, Transfection, Flow Cytometry, Cell Culture, Infection

2) Product Images from "ADP-ribose polymers localized on Ctcf-Parp1-Dnmt1 complex prevent methylation of Ctcf target sites"

Article Title: ADP-ribose polymers localized on Ctcf-Parp1-Dnmt1 complex prevent methylation of Ctcf target sites

Journal: Biochemical Journal

doi: 10.1042/BJ20111417

ChIP analysis of DMR1 occupancy by Ctcf, Parp1, PARs and Dnmt1 ( A ) Schematic map of the locus. The DMR1 region is expanded to show the approximate position of putative Ctcf-binding sites and of PCR primers used to detect the presence of specific DNA sequences in ChIP complexes. Circles represent CpG dinuclotides. H, HpaII sites. ( B ) ChIP analysis of DMR1 region carried out with anti-Ctcf, anti-Parp1, anti-Dnmt1 and anti-PAR Abs. Beads alone (No Ab) and anti-IgG Abs were used as negative controls. Real-time PCR data are expressed as percentage of the signal detected for the non-immunoprecipitated input (4% of the chromatin subjected to immunoprecipitation) taken as 100%. The Actb promoter served as a control. ( C ) Standard ChIP assay for Ctcf was followed by RE-ChIP to assess the co-occupancy of Ctcf, with Parp1, PARs and Dnmt1 at the Ctcf DNA target sites within the DMR1. Real-time PCR was performed for the DMR1 b and c fragments. The efficiency of RE-ChIP at each of the sites was calculated as a percentage of the chromatin input that co-purified with Ctcf in the first ChIP (10% of the Ctcf ChIP fraction). Results are means±S.E.M. calculated from three experiments. * P
Figure Legend Snippet: ChIP analysis of DMR1 occupancy by Ctcf, Parp1, PARs and Dnmt1 ( A ) Schematic map of the locus. The DMR1 region is expanded to show the approximate position of putative Ctcf-binding sites and of PCR primers used to detect the presence of specific DNA sequences in ChIP complexes. Circles represent CpG dinuclotides. H, HpaII sites. ( B ) ChIP analysis of DMR1 region carried out with anti-Ctcf, anti-Parp1, anti-Dnmt1 and anti-PAR Abs. Beads alone (No Ab) and anti-IgG Abs were used as negative controls. Real-time PCR data are expressed as percentage of the signal detected for the non-immunoprecipitated input (4% of the chromatin subjected to immunoprecipitation) taken as 100%. The Actb promoter served as a control. ( C ) Standard ChIP assay for Ctcf was followed by RE-ChIP to assess the co-occupancy of Ctcf, with Parp1, PARs and Dnmt1 at the Ctcf DNA target sites within the DMR1. Real-time PCR was performed for the DMR1 b and c fragments. The efficiency of RE-ChIP at each of the sites was calculated as a percentage of the chromatin input that co-purified with Ctcf in the first ChIP (10% of the Ctcf ChIP fraction). Results are means±S.E.M. calculated from three experiments. * P

Techniques Used: Chromatin Immunoprecipitation, Binding Assay, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Immunoprecipitation, Purification

Ctcf, Parp1, and Dnmt1 bind to non-methylated DNA molecules within the DMR1 Ctcf, Parp1, and Dnmt1 ChIPs were performed for the DMR1 b and c fragments. End-point PCR was performed after digestion of ChIP fractions DNA and inputs with either HpaII (H, methylation sensitive) or MspI (M, methylation insensitive) following heat inactivation of the restriction enzymes. The input (4% of the chromatin subjected to immunoprecipitation) was diluted 1/100 or 1/200 before restriction. The uncut (U) fractions consisted in HpaII digestions preventively blocked by heat inactivation. W, PCR performed in the absence of added template.
Figure Legend Snippet: Ctcf, Parp1, and Dnmt1 bind to non-methylated DNA molecules within the DMR1 Ctcf, Parp1, and Dnmt1 ChIPs were performed for the DMR1 b and c fragments. End-point PCR was performed after digestion of ChIP fractions DNA and inputs with either HpaII (H, methylation sensitive) or MspI (M, methylation insensitive) following heat inactivation of the restriction enzymes. The input (4% of the chromatin subjected to immunoprecipitation) was diluted 1/100 or 1/200 before restriction. The uncut (U) fractions consisted in HpaII digestions preventively blocked by heat inactivation. W, PCR performed in the absence of added template.

Techniques Used: Methylation, Polymerase Chain Reaction, Chromatin Immunoprecipitation, Immunoprecipitation

3) Product Images from "Novel BRCA1 and BRCA2 Tumor Test as Basis for Treatment Decisions and Referral for Genetic Counselling of Patients with Ovarian Carcinomas"

Article Title: Novel BRCA1 and BRCA2 Tumor Test as Basis for Treatment Decisions and Referral for Genetic Counselling of Patients with Ovarian Carcinomas

Journal: Human Mutation

doi: 10.1002/humu.23137

A : Pathogenic germline and somatic mutations in BRCA1 and BRCA2 detected using smMIP‐based targeted sequencing of FFPE tumor material. Lollipops above the bar: germline mutations detected in 47 FFPE ovarian carcinomas derived from 38 patients. Lollipops below the bar: somatic pathogenic mutations observed in seven FFPE ovarian carcinomas derived from five patients. B : Genomic location of 43 germline SNPs in BRCA1 and BRCA2 that were selected to determine the sensitivity of smMIP‐based next‐generation sequencing. Depicted base substitutions (lollipops) represent 15 and 28 benign germline variants in the ORF of BRCA1 and BRCA2 , respectively. These variants were known to be present in the germline of a subset of the included ovarian carcinoma patients prior to smMIP‐based sequencing of BRCA1 and BRCA2 in the corresponding FFPE ovarian carcinomas. Numbers depict the total number of the corresponding germline variant observed in these ovarian carcinoma patients. All germline SNPs could successfully be detected using smMIP‐based targeted sequencing on DNA derived from the corresponding FFPE ovarian carcinoma sample.
Figure Legend Snippet: A : Pathogenic germline and somatic mutations in BRCA1 and BRCA2 detected using smMIP‐based targeted sequencing of FFPE tumor material. Lollipops above the bar: germline mutations detected in 47 FFPE ovarian carcinomas derived from 38 patients. Lollipops below the bar: somatic pathogenic mutations observed in seven FFPE ovarian carcinomas derived from five patients. B : Genomic location of 43 germline SNPs in BRCA1 and BRCA2 that were selected to determine the sensitivity of smMIP‐based next‐generation sequencing. Depicted base substitutions (lollipops) represent 15 and 28 benign germline variants in the ORF of BRCA1 and BRCA2 , respectively. These variants were known to be present in the germline of a subset of the included ovarian carcinoma patients prior to smMIP‐based sequencing of BRCA1 and BRCA2 in the corresponding FFPE ovarian carcinomas. Numbers depict the total number of the corresponding germline variant observed in these ovarian carcinoma patients. All germline SNPs could successfully be detected using smMIP‐based targeted sequencing on DNA derived from the corresponding FFPE ovarian carcinoma sample.

Techniques Used: Sequencing, Formalin-fixed Paraffin-Embedded, Derivative Assay, Next-Generation Sequencing, Variant Assay

Related Articles

CRISPR:

Article Title: Activation of the miR-371/372/373 miRNA Cluster Enhances Oncogenicity and Drug Resistance in Oral Carcinoma Cells
Article Snippet: TransFectinTM Lipid Reagent (BioRad, Hercules, CA, USA) was used for transfection. .. Establishment of the CRISPR/Cas9 System Targeting Deletion of the miR-371/372/373 Cluster and the miR-371/372/373 Promoter The sequences spanning miR-371/372/373 cluster and the miR-371/372/373 promoter region were sent to the ATUM ( https://www.atum.bio/eCommerce/cas9/input ) portal in order to design small oligonucleotides that would guide the Cas9 complex and allow it to cleave the downstream sequence of the PAM (Protospacer Adjacent Motif, NGG) site. .. The oligunocleotides ( ) containing the guide sequences were annealed to form double stranded DNA and cloned into All-in-one pSpCas9-BB-2A-GFP-PX458 vector (Cat No.: 48138, Addgene, Cambridge, MA, USA) after cleavage by the BbsI restriction enzyme.

Sequencing:

Article Title: Activation of the miR-371/372/373 miRNA Cluster Enhances Oncogenicity and Drug Resistance in Oral Carcinoma Cells
Article Snippet: TransFectinTM Lipid Reagent (BioRad, Hercules, CA, USA) was used for transfection. .. Establishment of the CRISPR/Cas9 System Targeting Deletion of the miR-371/372/373 Cluster and the miR-371/372/373 Promoter The sequences spanning miR-371/372/373 cluster and the miR-371/372/373 promoter region were sent to the ATUM ( https://www.atum.bio/eCommerce/cas9/input ) portal in order to design small oligonucleotides that would guide the Cas9 complex and allow it to cleave the downstream sequence of the PAM (Protospacer Adjacent Motif, NGG) site. .. The oligunocleotides ( ) containing the guide sequences were annealed to form double stranded DNA and cloned into All-in-one pSpCas9-BB-2A-GFP-PX458 vector (Cat No.: 48138, Addgene, Cambridge, MA, USA) after cleavage by the BbsI restriction enzyme.

Amplification:

Article Title: Runx proteins mediate protective immunity against Leishmania donovani infection by promoting CD40 expression on dendritic cells
Article Snippet: The recruitment of Runx proteins to the mouse GAPDH promoter was examined by ChIP using indicated antibodies (left margin) and the primers described in “Materials and methods”. .. Amplification of chromatin immunoprecipitated by rabbit IgG served as a negative control, and input DNA (2%) as an internal control. ..

Immunoprecipitation:

Article Title: Runx proteins mediate protective immunity against Leishmania donovani infection by promoting CD40 expression on dendritic cells
Article Snippet: The recruitment of Runx proteins to the mouse GAPDH promoter was examined by ChIP using indicated antibodies (left margin) and the primers described in “Materials and methods”. .. Amplification of chromatin immunoprecipitated by rabbit IgG served as a negative control, and input DNA (2%) as an internal control. ..

Negative Control:

Article Title: Runx proteins mediate protective immunity against Leishmania donovani infection by promoting CD40 expression on dendritic cells
Article Snippet: The recruitment of Runx proteins to the mouse GAPDH promoter was examined by ChIP using indicated antibodies (left margin) and the primers described in “Materials and methods”. .. Amplification of chromatin immunoprecipitated by rabbit IgG served as a negative control, and input DNA (2%) as an internal control. ..

Real-time Polymerase Chain Reaction:

Article Title: STAT3 Partly Inhibits Cell Proliferation via Direct Negative Regulation of FST Gene Expression
Article Snippet: The qPCR reaction volume was 20 μL including 1 μL of cDNA, 10 μL of 2 × SYBR Green PCR MasterMix (Roche Molecular Systems, United States), 0.5 μL each of the forward and reverse primers (10 μM), and 8 μL double-distilled water. .. The qPCR conditions were as follows: 95°C for 10 min; 40 cycles at 95°C for 30 s, 60°C for 30 s. Non-immunoprecipitated DNA (2%) was used as input control. .. Two additional negative controls, mouse IgG (A) and anti-Myc antibody (B), were prepared by the co-transfection of HEK293T cells with pGL3-FST(−980/−340) and pCMV-Myc.

Article Title: Red Blood Cell Size Is Inversely Associated with Leukocyte Telomere Length in a Large Multi-Ethnic Population
Article Snippet: .. Here we have used a quantitative PCR (qPCR) method for estimation of length of telomeres relative to a single copy gene that requires only a small amount of input genomic DNA (20 ng per reaction). .. It is a limitation of the PCR-based nature of this assay that measurements of telomere size may not be completely accurate, which would affect our estimate of telomere attrition in basepairs.

Hybridization:

Article Title: A label-free and enzyme-free platform with a visible output for constructing versatile logic gates using caged G-quadruplex as the signal transducer label-free and enzyme-free platform with a visible output for constructing versatile logic gates using caged G-quadruplex as the signal transducer †Electronic supplementa
Article Snippet: .. In the presence of both inputs, the six DNAzyme subunits (DNA2–DNA7) cannot assemble into any active DNAzyme structure due to the preferred inter-input hybridization and the displacement reactions between the template and the inputs. .. Thus, parallel activation of a combinatorial circuit established from the XOR and NOR gates could be realized in a single system using the same set of inputs. shows typical photographs of the XOR + NOR gate. depicts the absorption spectra from 500 to 800 nm. shows the corresponding absorption intensity at λ = 650 nm.

Sonication:

Article Title: Tip60 functions as a potential corepressor of KLF4 in regulation of HDC promoter activity
Article Snippet: CHIPs AGSE cells in 10 cm dishes with and without 10−8 M gastrin treatment were fixed with 1% formaldehyde at 37°C for 10 min and then harvested and lysed with 500 µl of lysis buffer [1%SDS, 5 mM EDTA, 50 mM Tris/HCl (pH 8.0), plus protease inhibitor]. .. After brief sonication (3 times at 10 s each), extracts were precleared by 2 µg of sheared salmon sperm DNA, 20 µl of control IgG and protein A-Sepharose for 2 h at 4°C. ..

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    ATUM input dna
    LPS and TNFα induce nuclear localization and binding of Runx proteins to the CD40 promoter. (A) Schematic presentation of mouse CD40 promoter indicating the location of putative Runx-binding sites (R1 and R2) and ChIP primers (P1 and P2), and details of oligonucleotide probes used for EMSA. Mouse CD40 promoter-specific Pr1 and Pr2 probes contain putative wild-type Runx-binding sites, and Mut-Pr1 and Mut-Pr2 probes contain mutations (in italics) at Runx-binding sites. Base positions are relative to the transcription start site. (B and C) EMSA of nuclear extracts of BMDCs treated with LPS or TNFα for indicated times; assessed with indicated probes. Numbers below lanes in (B) represent densitometry [normalized to OCT-1 binding (control)] relative to that of untreated BMDCs (0 h). The densitometry results pooled from three independent experiments for (B) are shown in S2 Fig . (D) Supershift EMSA [with immunoglobulin G (IgG; control) or antibody to (α-) Runx1, Runx2 or Runx3] of nuclear extracts of BMDCs treated with LPS or TNFα for 0.5 h; assayed with Pr1 or Pr2 probes. (E) Binding of Runx proteins to -578/-373 region of the CD40 promoter in BMDCs treated with LPS or TNFα (time, above lanes); assessed via ChIP using the primers shown in (A) and indicated antibodies (left margin). Amplification of mouse GAPDH promoter ( S4 Fig ) and chromatin <t>immunoprecipitated</t> by rabbit IgG were used as negative controls, and input <t>DNA</t> (2%) as an internal control. Numbers below lanes represent densitometry, normalized to input DNA and presented relative to that of untreated BMDCs (0 h). (F) In vivo footprint analysis of mouse CD40 promoter showing DMS-protected bands (indicated by open circles) at R1 and R2 sites in LPS- or TNFα-treated but not in untreated (UT) BMDCs. (G) Translocation of Runx1 or Runx3 (red) into the nuclei (blue; Hoechst staining) in BMDCs left untreated (UT) or treated with LPS or TNFα for 0.5 h; assessed by confocal microscopy. Pink color (merge) shows nuclear translocation of Runx1 or Runx3. Scale bar, 10 μm. Confocal microscopy images of DCs immunostained with isotype control antibodies are given in S5 Fig . Data are representative of three (B-E) or two (F and G) independent experiments.
    Input Dna, supplied by ATUM, 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/input dna/product/ATUM
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    input dna - by Bioz Stars, 2021-07
    86/100 stars
      Buy from Supplier

    86
    ATUM minion low input library preparation kit
    Function and Taxonomy of Ice Wedge soil. Ice wedge soil metagenomes sequenced using the <t>MinION</t> rapid and <t>low</t> <t>input</t> <t>kit</t> (A) Taxonomy at the domain level (B) Clusters of Orthologous Groups (COG) categories (level 2).
    Minion Low Input Library Preparation Kit, supplied by ATUM, 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/minion low input library preparation kit/product/ATUM
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    minion low input library preparation kit - by Bioz Stars, 2021-07
    86/100 stars
      Buy from Supplier

    Image Search Results


    LPS and TNFα induce nuclear localization and binding of Runx proteins to the CD40 promoter. (A) Schematic presentation of mouse CD40 promoter indicating the location of putative Runx-binding sites (R1 and R2) and ChIP primers (P1 and P2), and details of oligonucleotide probes used for EMSA. Mouse CD40 promoter-specific Pr1 and Pr2 probes contain putative wild-type Runx-binding sites, and Mut-Pr1 and Mut-Pr2 probes contain mutations (in italics) at Runx-binding sites. Base positions are relative to the transcription start site. (B and C) EMSA of nuclear extracts of BMDCs treated with LPS or TNFα for indicated times; assessed with indicated probes. Numbers below lanes in (B) represent densitometry [normalized to OCT-1 binding (control)] relative to that of untreated BMDCs (0 h). The densitometry results pooled from three independent experiments for (B) are shown in S2 Fig . (D) Supershift EMSA [with immunoglobulin G (IgG; control) or antibody to (α-) Runx1, Runx2 or Runx3] of nuclear extracts of BMDCs treated with LPS or TNFα for 0.5 h; assayed with Pr1 or Pr2 probes. (E) Binding of Runx proteins to -578/-373 region of the CD40 promoter in BMDCs treated with LPS or TNFα (time, above lanes); assessed via ChIP using the primers shown in (A) and indicated antibodies (left margin). Amplification of mouse GAPDH promoter ( S4 Fig ) and chromatin immunoprecipitated by rabbit IgG were used as negative controls, and input DNA (2%) as an internal control. Numbers below lanes represent densitometry, normalized to input DNA and presented relative to that of untreated BMDCs (0 h). (F) In vivo footprint analysis of mouse CD40 promoter showing DMS-protected bands (indicated by open circles) at R1 and R2 sites in LPS- or TNFα-treated but not in untreated (UT) BMDCs. (G) Translocation of Runx1 or Runx3 (red) into the nuclei (blue; Hoechst staining) in BMDCs left untreated (UT) or treated with LPS or TNFα for 0.5 h; assessed by confocal microscopy. Pink color (merge) shows nuclear translocation of Runx1 or Runx3. Scale bar, 10 μm. Confocal microscopy images of DCs immunostained with isotype control antibodies are given in S5 Fig . Data are representative of three (B-E) or two (F and G) independent experiments.

    Journal: PLoS Pathogens

    Article Title: Runx proteins mediate protective immunity against Leishmania donovani infection by promoting CD40 expression on dendritic cells

    doi: 10.1371/journal.ppat.1009136

    Figure Lengend Snippet: LPS and TNFα induce nuclear localization and binding of Runx proteins to the CD40 promoter. (A) Schematic presentation of mouse CD40 promoter indicating the location of putative Runx-binding sites (R1 and R2) and ChIP primers (P1 and P2), and details of oligonucleotide probes used for EMSA. Mouse CD40 promoter-specific Pr1 and Pr2 probes contain putative wild-type Runx-binding sites, and Mut-Pr1 and Mut-Pr2 probes contain mutations (in italics) at Runx-binding sites. Base positions are relative to the transcription start site. (B and C) EMSA of nuclear extracts of BMDCs treated with LPS or TNFα for indicated times; assessed with indicated probes. Numbers below lanes in (B) represent densitometry [normalized to OCT-1 binding (control)] relative to that of untreated BMDCs (0 h). The densitometry results pooled from three independent experiments for (B) are shown in S2 Fig . (D) Supershift EMSA [with immunoglobulin G (IgG; control) or antibody to (α-) Runx1, Runx2 or Runx3] of nuclear extracts of BMDCs treated with LPS or TNFα for 0.5 h; assayed with Pr1 or Pr2 probes. (E) Binding of Runx proteins to -578/-373 region of the CD40 promoter in BMDCs treated with LPS or TNFα (time, above lanes); assessed via ChIP using the primers shown in (A) and indicated antibodies (left margin). Amplification of mouse GAPDH promoter ( S4 Fig ) and chromatin immunoprecipitated by rabbit IgG were used as negative controls, and input DNA (2%) as an internal control. Numbers below lanes represent densitometry, normalized to input DNA and presented relative to that of untreated BMDCs (0 h). (F) In vivo footprint analysis of mouse CD40 promoter showing DMS-protected bands (indicated by open circles) at R1 and R2 sites in LPS- or TNFα-treated but not in untreated (UT) BMDCs. (G) Translocation of Runx1 or Runx3 (red) into the nuclei (blue; Hoechst staining) in BMDCs left untreated (UT) or treated with LPS or TNFα for 0.5 h; assessed by confocal microscopy. Pink color (merge) shows nuclear translocation of Runx1 or Runx3. Scale bar, 10 μm. Confocal microscopy images of DCs immunostained with isotype control antibodies are given in S5 Fig . Data are representative of three (B-E) or two (F and G) independent experiments.

    Article Snippet: Amplification of chromatin immunoprecipitated by rabbit IgG served as a negative control, and input DNA (2%) as an internal control.

    Techniques: Binding Assay, Chromatin Immunoprecipitation, Amplification, Immunoprecipitation, In Vivo, Translocation Assay, Staining, Confocal Microscopy

    Sb R LD inhibits SAG-induced CD40 upregulation on HuMoDCs by impairing Runx binding to the CD40 promoter. (A) A schematic of human CD40 promoter showing the location of putative Runx-binding sites (R3 and R4) and ChIP primers (P3 and P4), and details of oligonucleotide probes used for EMSA. Human CD40 promoter-specific Pr3 and Pr4 probes contain putative wild-type Runx-binding sites, and Mut-Pr3 and Mut-Pr4 probes contain mutations (in italics) at Runx-binding sites. Base positions are relative to the transcription start site. (B and C) EMSA of nuclear extracts of HuMoDCs treated with SAG for indicated times; assayed with indicated probes. OCT-1 binding (in B) serves as an internal control. Numbers below lanes in (B), densitometry readings (as in Fig 2B ); presented relative to untreated HuMoDCs (0 h). (D) Supershift EMSA (antibodies and probes are indicated above and below lanes, respectively) to assess the binding of nuclear Runx1, Runx2 and Runx3 to the CD40 promoter in HuMoDCs treated with SAG for 0.5 h. (E) ChIP assay [with the primers shown in (A) and antibodies indicated at left margin] to determine the binding of Runx proteins to -480/-276 region of the CD40 promoter in HuMoDCs treated with SAG (time, above lanes). Amplification of human GAPDH promoter and chromatin immunoprecipitated by rabbit IgG were used as negative controls, and input DNA (2%) as an internal control. Numbers below lanes represent densitometry, normalized to input DNA and presented relative to that of untreated HuMoDCs (0 h). hu CD40 , human CD40 ; hu GAPDH , human GAPDH . (F) Immunoblot analysis of Runx1 and Runx3 expression in HuMoDCs left untransfected, or transfected with control siRNA, Runx1 siRNA, Runx3 siRNA or Runx1 and Runx3 siRNAs (Runx1+3 siRNA). β-actin serves as a loading control. (G) Flow cytometry analysis of CD40 expression on HuMoDCs transfected with indicated siRNAs and then cultured with or without SAG for 24 h. (H) EMSA of nuclear extracts of HuMoDCs left uninfected or infected with promastigotes of Sb R LD strain GE1F8R or Sb S LD strain AG83 for 24 h and then cultured for 0.5 h with (+) or without (-) SAG; assayed with indicated probes. OCT-1 binding serves as an internal control. Numbers below lanes represent densitometry readings (as in Fig 2B ); presented relative to control HuMoDCs (HuMoDCs left uninfected and cultured without SAG). (I) HuMoDCs were either left uninfected or infected with promastigotes of Sb S LD strain AG83 or Sb R LD strain GE1F8R for 24 h and then treated (for 24 h) with or without SAG. CD40 expression on HuMoDCs was assessed by flow cytometry. All data are representative of two independent experiments.

    Journal: PLoS Pathogens

    Article Title: Runx proteins mediate protective immunity against Leishmania donovani infection by promoting CD40 expression on dendritic cells

    doi: 10.1371/journal.ppat.1009136

    Figure Lengend Snippet: Sb R LD inhibits SAG-induced CD40 upregulation on HuMoDCs by impairing Runx binding to the CD40 promoter. (A) A schematic of human CD40 promoter showing the location of putative Runx-binding sites (R3 and R4) and ChIP primers (P3 and P4), and details of oligonucleotide probes used for EMSA. Human CD40 promoter-specific Pr3 and Pr4 probes contain putative wild-type Runx-binding sites, and Mut-Pr3 and Mut-Pr4 probes contain mutations (in italics) at Runx-binding sites. Base positions are relative to the transcription start site. (B and C) EMSA of nuclear extracts of HuMoDCs treated with SAG for indicated times; assayed with indicated probes. OCT-1 binding (in B) serves as an internal control. Numbers below lanes in (B), densitometry readings (as in Fig 2B ); presented relative to untreated HuMoDCs (0 h). (D) Supershift EMSA (antibodies and probes are indicated above and below lanes, respectively) to assess the binding of nuclear Runx1, Runx2 and Runx3 to the CD40 promoter in HuMoDCs treated with SAG for 0.5 h. (E) ChIP assay [with the primers shown in (A) and antibodies indicated at left margin] to determine the binding of Runx proteins to -480/-276 region of the CD40 promoter in HuMoDCs treated with SAG (time, above lanes). Amplification of human GAPDH promoter and chromatin immunoprecipitated by rabbit IgG were used as negative controls, and input DNA (2%) as an internal control. Numbers below lanes represent densitometry, normalized to input DNA and presented relative to that of untreated HuMoDCs (0 h). hu CD40 , human CD40 ; hu GAPDH , human GAPDH . (F) Immunoblot analysis of Runx1 and Runx3 expression in HuMoDCs left untransfected, or transfected with control siRNA, Runx1 siRNA, Runx3 siRNA or Runx1 and Runx3 siRNAs (Runx1+3 siRNA). β-actin serves as a loading control. (G) Flow cytometry analysis of CD40 expression on HuMoDCs transfected with indicated siRNAs and then cultured with or without SAG for 24 h. (H) EMSA of nuclear extracts of HuMoDCs left uninfected or infected with promastigotes of Sb R LD strain GE1F8R or Sb S LD strain AG83 for 24 h and then cultured for 0.5 h with (+) or without (-) SAG; assayed with indicated probes. OCT-1 binding serves as an internal control. Numbers below lanes represent densitometry readings (as in Fig 2B ); presented relative to control HuMoDCs (HuMoDCs left uninfected and cultured without SAG). (I) HuMoDCs were either left uninfected or infected with promastigotes of Sb S LD strain AG83 or Sb R LD strain GE1F8R for 24 h and then treated (for 24 h) with or without SAG. CD40 expression on HuMoDCs was assessed by flow cytometry. All data are representative of two independent experiments.

    Article Snippet: Amplification of chromatin immunoprecipitated by rabbit IgG served as a negative control, and input DNA (2%) as an internal control.

    Techniques: Binding Assay, Chromatin Immunoprecipitation, Amplification, Immunoprecipitation, Expressing, Transfection, Flow Cytometry, Cell Culture, Infection

    ChIP analysis of DMR1 occupancy by Ctcf, Parp1, PARs and Dnmt1 ( A ) Schematic map of the locus. The DMR1 region is expanded to show the approximate position of putative Ctcf-binding sites and of PCR primers used to detect the presence of specific DNA sequences in ChIP complexes. Circles represent CpG dinuclotides. H, HpaII sites. ( B ) ChIP analysis of DMR1 region carried out with anti-Ctcf, anti-Parp1, anti-Dnmt1 and anti-PAR Abs. Beads alone (No Ab) and anti-IgG Abs were used as negative controls. Real-time PCR data are expressed as percentage of the signal detected for the non-immunoprecipitated input (4% of the chromatin subjected to immunoprecipitation) taken as 100%. The Actb promoter served as a control. ( C ) Standard ChIP assay for Ctcf was followed by RE-ChIP to assess the co-occupancy of Ctcf, with Parp1, PARs and Dnmt1 at the Ctcf DNA target sites within the DMR1. Real-time PCR was performed for the DMR1 b and c fragments. The efficiency of RE-ChIP at each of the sites was calculated as a percentage of the chromatin input that co-purified with Ctcf in the first ChIP (10% of the Ctcf ChIP fraction). Results are means±S.E.M. calculated from three experiments. * P

    Journal: Biochemical Journal

    Article Title: ADP-ribose polymers localized on Ctcf-Parp1-Dnmt1 complex prevent methylation of Ctcf target sites

    doi: 10.1042/BJ20111417

    Figure Lengend Snippet: ChIP analysis of DMR1 occupancy by Ctcf, Parp1, PARs and Dnmt1 ( A ) Schematic map of the locus. The DMR1 region is expanded to show the approximate position of putative Ctcf-binding sites and of PCR primers used to detect the presence of specific DNA sequences in ChIP complexes. Circles represent CpG dinuclotides. H, HpaII sites. ( B ) ChIP analysis of DMR1 region carried out with anti-Ctcf, anti-Parp1, anti-Dnmt1 and anti-PAR Abs. Beads alone (No Ab) and anti-IgG Abs were used as negative controls. Real-time PCR data are expressed as percentage of the signal detected for the non-immunoprecipitated input (4% of the chromatin subjected to immunoprecipitation) taken as 100%. The Actb promoter served as a control. ( C ) Standard ChIP assay for Ctcf was followed by RE-ChIP to assess the co-occupancy of Ctcf, with Parp1, PARs and Dnmt1 at the Ctcf DNA target sites within the DMR1. Real-time PCR was performed for the DMR1 b and c fragments. The efficiency of RE-ChIP at each of the sites was calculated as a percentage of the chromatin input that co-purified with Ctcf in the first ChIP (10% of the Ctcf ChIP fraction). Results are means±S.E.M. calculated from three experiments. * P

    Article Snippet: ChIP-CHOP assay For ChIP-CHOP analysis of the DMR1 b and c fragments, the Ctcf-, Parp1- and Dnmt-pulled down DNA (20 μl), and input DNA (20 μl after 1:100 and 1:200 dilutions) from ChIP assays were digested for 1 h at 37°C with 10 units of either the methylation-sensitive HpaII restriction enzyme or its methylation-insensitive isoschizomer MspI (New England Biolabs).

    Techniques: Chromatin Immunoprecipitation, Binding Assay, Polymerase Chain Reaction, Real-time Polymerase Chain Reaction, Immunoprecipitation, Purification

    Ctcf, Parp1, and Dnmt1 bind to non-methylated DNA molecules within the DMR1 Ctcf, Parp1, and Dnmt1 ChIPs were performed for the DMR1 b and c fragments. End-point PCR was performed after digestion of ChIP fractions DNA and inputs with either HpaII (H, methylation sensitive) or MspI (M, methylation insensitive) following heat inactivation of the restriction enzymes. The input (4% of the chromatin subjected to immunoprecipitation) was diluted 1/100 or 1/200 before restriction. The uncut (U) fractions consisted in HpaII digestions preventively blocked by heat inactivation. W, PCR performed in the absence of added template.

    Journal: Biochemical Journal

    Article Title: ADP-ribose polymers localized on Ctcf-Parp1-Dnmt1 complex prevent methylation of Ctcf target sites

    doi: 10.1042/BJ20111417

    Figure Lengend Snippet: Ctcf, Parp1, and Dnmt1 bind to non-methylated DNA molecules within the DMR1 Ctcf, Parp1, and Dnmt1 ChIPs were performed for the DMR1 b and c fragments. End-point PCR was performed after digestion of ChIP fractions DNA and inputs with either HpaII (H, methylation sensitive) or MspI (M, methylation insensitive) following heat inactivation of the restriction enzymes. The input (4% of the chromatin subjected to immunoprecipitation) was diluted 1/100 or 1/200 before restriction. The uncut (U) fractions consisted in HpaII digestions preventively blocked by heat inactivation. W, PCR performed in the absence of added template.

    Article Snippet: ChIP-CHOP assay For ChIP-CHOP analysis of the DMR1 b and c fragments, the Ctcf-, Parp1- and Dnmt-pulled down DNA (20 μl), and input DNA (20 μl after 1:100 and 1:200 dilutions) from ChIP assays were digested for 1 h at 37°C with 10 units of either the methylation-sensitive HpaII restriction enzyme or its methylation-insensitive isoschizomer MspI (New England Biolabs).

    Techniques: Methylation, Polymerase Chain Reaction, Chromatin Immunoprecipitation, Immunoprecipitation

    A : Pathogenic germline and somatic mutations in BRCA1 and BRCA2 detected using smMIP‐based targeted sequencing of FFPE tumor material. Lollipops above the bar: germline mutations detected in 47 FFPE ovarian carcinomas derived from 38 patients. Lollipops below the bar: somatic pathogenic mutations observed in seven FFPE ovarian carcinomas derived from five patients. B : Genomic location of 43 germline SNPs in BRCA1 and BRCA2 that were selected to determine the sensitivity of smMIP‐based next‐generation sequencing. Depicted base substitutions (lollipops) represent 15 and 28 benign germline variants in the ORF of BRCA1 and BRCA2 , respectively. These variants were known to be present in the germline of a subset of the included ovarian carcinoma patients prior to smMIP‐based sequencing of BRCA1 and BRCA2 in the corresponding FFPE ovarian carcinomas. Numbers depict the total number of the corresponding germline variant observed in these ovarian carcinoma patients. All germline SNPs could successfully be detected using smMIP‐based targeted sequencing on DNA derived from the corresponding FFPE ovarian carcinoma sample.

    Journal: Human Mutation

    Article Title: Novel BRCA1 and BRCA2 Tumor Test as Basis for Treatment Decisions and Referral for Genetic Counselling of Patients with Ovarian Carcinomas

    doi: 10.1002/humu.23137

    Figure Lengend Snippet: A : Pathogenic germline and somatic mutations in BRCA1 and BRCA2 detected using smMIP‐based targeted sequencing of FFPE tumor material. Lollipops above the bar: germline mutations detected in 47 FFPE ovarian carcinomas derived from 38 patients. Lollipops below the bar: somatic pathogenic mutations observed in seven FFPE ovarian carcinomas derived from five patients. B : Genomic location of 43 germline SNPs in BRCA1 and BRCA2 that were selected to determine the sensitivity of smMIP‐based next‐generation sequencing. Depicted base substitutions (lollipops) represent 15 and 28 benign germline variants in the ORF of BRCA1 and BRCA2 , respectively. These variants were known to be present in the germline of a subset of the included ovarian carcinoma patients prior to smMIP‐based sequencing of BRCA1 and BRCA2 in the corresponding FFPE ovarian carcinomas. Numbers depict the total number of the corresponding germline variant observed in these ovarian carcinoma patients. All germline SNPs could successfully be detected using smMIP‐based targeted sequencing on DNA derived from the corresponding FFPE ovarian carcinoma sample.

    Article Snippet: Briefly, smMIP capture was performed on 10μl of input DNA (20–500ng) supplied with 15μl capture mixture (0.01μl ampligase DNA ligase [100U/μl; Illumina, Madison, WI], 2.5μl 10x ampligase buffer [Illumina], 0.27μl smMIP pool dilution [6.6x105 μM], 0.32μl Hemo Klentaq [10U/μl; New England Biolabs], 0.03μl dNTPs [0.25mM], and 11.9μl H2 O).

    Techniques: Sequencing, Formalin-fixed Paraffin-Embedded, Derivative Assay, Next-Generation Sequencing, Variant Assay

    Function and Taxonomy of Ice Wedge soil. Ice wedge soil metagenomes sequenced using the MinION rapid and low input kit (A) Taxonomy at the domain level (B) Clusters of Orthologous Groups (COG) categories (level 2).

    Journal: Frontiers in Microbiology

    Article Title: In Situ Field Sequencing and Life Detection in Remote (79°26′N) Canadian High Arctic Permafrost Ice Wedge Microbial Communities

    doi: 10.3389/fmicb.2017.02594

    Figure Lengend Snippet: Function and Taxonomy of Ice Wedge soil. Ice wedge soil metagenomes sequenced using the MinION rapid and low input kit (A) Taxonomy at the domain level (B) Clusters of Orthologous Groups (COG) categories (level 2).

    Article Snippet: The MinION low input library preparation kit required much lower amounts of DNA (~20–100 ng), took ~160 min and required more instrumentation (a microcentrifuge, magnetic beads, additional enzymes, ligation of a hairpin adapter etc.)

    Techniques:

    Ice Wedge soil bacterial community composition detected by the MinION. Bacterial reads from ice wedge soil metagenomes sequenced using the MinION rapid and low input kit compared with Bacterial composition inferred from amplicon sequencing of 16S rRNA gene.

    Journal: Frontiers in Microbiology

    Article Title: In Situ Field Sequencing and Life Detection in Remote (79°26′N) Canadian High Arctic Permafrost Ice Wedge Microbial Communities

    doi: 10.3389/fmicb.2017.02594

    Figure Lengend Snippet: Ice Wedge soil bacterial community composition detected by the MinION. Bacterial reads from ice wedge soil metagenomes sequenced using the MinION rapid and low input kit compared with Bacterial composition inferred from amplicon sequencing of 16S rRNA gene.

    Article Snippet: The MinION low input library preparation kit required much lower amounts of DNA (~20–100 ng), took ~160 min and required more instrumentation (a microcentrifuge, magnetic beads, additional enzymes, ligation of a hairpin adapter etc.)

    Techniques: Amplification, Sequencing