Structured Review

TaKaRa genome walker pcr
RNA-Sequencing analysis of <t>GhABF2</t> -overexpressing cotton leaves transcriptome. ( a ) Changes in gene expression profile between control, OE17, and OE18. DEGs, differently expressed genes. vs, versus. The red bar represents up-regulated gene, and the green bar represents down-regulated gene. ( b ) Venn diagram of DEGs in cotton seedling leaves between OE17 vs OE18 and different abiotic stress conditions. The DEGs data response to ABA, drought, and salt treatment were extracted from GEO at NCBI (accession number GSE50770). ( c ) Heatmap of the part of 68 DEGs. TF, transcription factor. OR, oxidation reduction. CB, chlorophyll biosynthetic. ( d ) Expression of TF genes by <t>qRT-PCR.</t> Values are means ± SD of three replicates. * P ≤ 0.01, ** P ≤ 0.01; Student t test.
Genome Walker Pcr, supplied by TaKaRa, used in various techniques. Bioz Stars score: 85/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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1) Product Images from "GhABF2, a bZIP transcription factor, confers drought and salinity tolerance in cotton (Gossypium hirsutum L.)"

Article Title: GhABF2, a bZIP transcription factor, confers drought and salinity tolerance in cotton (Gossypium hirsutum L.)

Journal: Scientific Reports

doi: 10.1038/srep35040

RNA-Sequencing analysis of GhABF2 -overexpressing cotton leaves transcriptome. ( a ) Changes in gene expression profile between control, OE17, and OE18. DEGs, differently expressed genes. vs, versus. The red bar represents up-regulated gene, and the green bar represents down-regulated gene. ( b ) Venn diagram of DEGs in cotton seedling leaves between OE17 vs OE18 and different abiotic stress conditions. The DEGs data response to ABA, drought, and salt treatment were extracted from GEO at NCBI (accession number GSE50770). ( c ) Heatmap of the part of 68 DEGs. TF, transcription factor. OR, oxidation reduction. CB, chlorophyll biosynthetic. ( d ) Expression of TF genes by qRT-PCR. Values are means ± SD of three replicates. * P ≤ 0.01, ** P ≤ 0.01; Student t test.
Figure Legend Snippet: RNA-Sequencing analysis of GhABF2 -overexpressing cotton leaves transcriptome. ( a ) Changes in gene expression profile between control, OE17, and OE18. DEGs, differently expressed genes. vs, versus. The red bar represents up-regulated gene, and the green bar represents down-regulated gene. ( b ) Venn diagram of DEGs in cotton seedling leaves between OE17 vs OE18 and different abiotic stress conditions. The DEGs data response to ABA, drought, and salt treatment were extracted from GEO at NCBI (accession number GSE50770). ( c ) Heatmap of the part of 68 DEGs. TF, transcription factor. OR, oxidation reduction. CB, chlorophyll biosynthetic. ( d ) Expression of TF genes by qRT-PCR. Values are means ± SD of three replicates. * P ≤ 0.01, ** P ≤ 0.01; Student t test.

Techniques Used: RNA Sequencing Assay, Expressing, Quantitative RT-PCR

Related Articles

Sequencing:

Article Title: GhABF2, a bZIP transcription factor, confers drought and salinity tolerance in cotton (Gossypium hirsutum L.)
Article Snippet: .. The Genome sequence of GhABF2 was isolated by Genome Walker PCR (TaKaRa, Dalian, China). .. Full-length open reading fragment (ORF) of GhABF2 was isolated by Invitrogen RACE system.

Article Title: Regulation of Stomatal Tropism and Infection by Light in Cercospora zeae-maydis: Evidence for Coordinated Host/Pathogen Responses to Photoperiod?
Article Snippet: .. The remainder of CRP1 was obtained by genome-walker PCR (Clontech) and sequencing clones containing CRP1 identified in a cosmid library containing 8× coverage of the C. zeae-maydis genome (kindly provided by Dr. Won-Bo Shim, Texas A & M University). ..

Isolation:

Article Title: GhABF2, a bZIP transcription factor, confers drought and salinity tolerance in cotton (Gossypium hirsutum L.)
Article Snippet: .. The Genome sequence of GhABF2 was isolated by Genome Walker PCR (TaKaRa, Dalian, China). .. Full-length open reading fragment (ORF) of GhABF2 was isolated by Invitrogen RACE system.

Polymerase Chain Reaction:

Article Title: GhABF2, a bZIP transcription factor, confers drought and salinity tolerance in cotton (Gossypium hirsutum L.)
Article Snippet: .. The Genome sequence of GhABF2 was isolated by Genome Walker PCR (TaKaRa, Dalian, China). .. Full-length open reading fragment (ORF) of GhABF2 was isolated by Invitrogen RACE system.

Article Title: Regulation of Stomatal Tropism and Infection by Light in Cercospora zeae-maydis: Evidence for Coordinated Host/Pathogen Responses to Photoperiod?
Article Snippet: .. The remainder of CRP1 was obtained by genome-walker PCR (Clontech) and sequencing clones containing CRP1 identified in a cosmid library containing 8× coverage of the C. zeae-maydis genome (kindly provided by Dr. Won-Bo Shim, Texas A & M University). ..

Clone Assay:

Article Title: Regulation of Stomatal Tropism and Infection by Light in Cercospora zeae-maydis: Evidence for Coordinated Host/Pathogen Responses to Photoperiod?
Article Snippet: .. The remainder of CRP1 was obtained by genome-walker PCR (Clontech) and sequencing clones containing CRP1 identified in a cosmid library containing 8× coverage of the C. zeae-maydis genome (kindly provided by Dr. Won-Bo Shim, Texas A & M University). ..

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  • 85
    TaKaRa genome walker dna walking
    Downregulation of <t>NtCYS</t> induces precocious cell death in the basal cell lineage and seed abortion. (A) Reduced expression of NtCYS in RNAi lines, as measured by RT-qPCR. The expression level of NtCYS in the WT was set to l. (B) The frequency of the two-celled proembryos with PI-positive basal cells in WT and RNAi lines. Data represent the mean ± SE from five independent experiments, with 50 proembryos per line analysed in each experiment ( n = 250). (C) Cell viability in the two-celled proembryos from WT and RNAi lines stained with FDA and PI. Scale bars, 10 µm. (D) Nuclear <t>DNA</t> fragmentation in the two-celled proembryos from WT and RNAi lines stained with TUNEL. Scale bars, 10 µm. (E) The frequency of two-celled proembryos with TUNEL-positive basal cells in WT and RNAi lines. Data represent the mean ± SE from three independent experiments, with 30 proembryos per line analysed in each experiment ( n = 90). (F–K) Morphology of the apical (G, J) and basal (H, K) cells analysed by TEM in the two-celled proembryos from WT (F–H) and RNAi lines (I–K). Scale bars, 5 µm in (F) and (I), and 2 µm in (G, H, J and K). Ac, apical cell; Bc, basal cell; n, nucleus; v, vacuole; nm, nuclear membrane; cw, cell wall; pm, plasma membrane. (L) Seed abortion in RNAi lines. Asterisks indicate aborted seeds. Scale bars, 1 mm. (M) Aberrant cell division patterns in the apical cell lineage at the early stages of embryogenesis in RNAi lines. Scale bars, 10 µm. (N) The frequency of aborted seeds in WT and RNAi lines. Data represent the mean ± SE from three independent experiments, with 400 to 500 seeds per line analysed in each experiment. (O, P) Cathepsin-like (O) and caspase-like (P) activities in the two-celled proembryos from WT and NtCYS RNAi line L2-6. Data represent the mean ± SE, with 40 to 60 two-celled proembryos. ** indicates statistical difference compared to WT ( t -test, p
    Genome Walker Dna Walking, supplied by TaKaRa, used in various techniques. Bioz Stars score: 85/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    97
    TaKaRa human genomic dna
    Effect of LCA treatment in HuH-7 cells on nuclear protein levels of Nrf2 and Mafs and ARE nuclear binding activity by these proteins. Part A ) Western blot analysis. Nuclear proteins were isolated from HuH-7 cells treated with LCA (100 µM) for up to 24 hours (15 µg protein/lane). Membranes were stripped and re-probed with histone 3 to ensure equal loading. Representative blots from 3 separate experiments are shown and numbers below the blots represent densitometric values expressed as % of 0 time control. Part B ) EMSA and supershift analysis. HuH-7 cells were treated with LCA (100 µM) or vehicle control for 20 hours and antibodies to Nrf1, Nrf2, MafG and c-Maf were used to compare relative amount of binding of these transcription factors to ARE as described in Methods. Part C ) ChIP analysis. HuH-7 cells were treated with LCA (100 µM) or vehicle control for 20 hours, then processed for ChIP assay as described in Methods. <t>PCR</t> products from amplification of the ARE sites following immunoprecipitation with antisera against Nrf2, Nrf1, c-Maf, MafG and Histone 3 demonstrate LCA treatment led to decreased Nrf2 and Nrf1 binding to the ARE sites, while c-Maf and MafG binding increased. Input genomic <t>DNA</t> (gDNA input) was used as a positive control and a no antibody immunoprecipitation (no Ab) was used as a negative control.
    Human Genomic Dna, supplied by TaKaRa, used in various techniques. Bioz Stars score: 97/100, based on 129 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    TaKaRa pcr based genome walker kit
    The T <t>DNA</t> insert is located in the promoter region of PIF3 . The diagram depicts (from top to bottom) the position of the poc1 locus on chromosome 1; the sequenced BAC F14J9 containing this locus; the T DNA insertion site in the promoter region of the PIF3 gene of poc1 and its location with respect to the predicted neighboring genes CAD and a MYB -related gene ( MYB ); the structure of the PIF3 gene and the location of the T DNA insert; and the structure of the PIF3 protein. The protein coding regions within the PIF3 gene are shown as filled boxes, the untranslated regions as open boxes, the introns and flanking DNA as a line, and the putative TATA box as vertical lines. The identity and location of motifs within the PIF3 protein are shown: PAS, Per–Arnt–Sim-like domain; bHLH, basic helix–loop–helix domain; NLS, nuclear localization signal. The position of the probe used in quantitative <t>PCR</t> analysis of PIF3 transcript levels also is shown (Probe).
    Pcr Based Genome Walker Kit, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 4 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Downregulation of NtCYS induces precocious cell death in the basal cell lineage and seed abortion. (A) Reduced expression of NtCYS in RNAi lines, as measured by RT-qPCR. The expression level of NtCYS in the WT was set to l. (B) The frequency of the two-celled proembryos with PI-positive basal cells in WT and RNAi lines. Data represent the mean ± SE from five independent experiments, with 50 proembryos per line analysed in each experiment ( n = 250). (C) Cell viability in the two-celled proembryos from WT and RNAi lines stained with FDA and PI. Scale bars, 10 µm. (D) Nuclear DNA fragmentation in the two-celled proembryos from WT and RNAi lines stained with TUNEL. Scale bars, 10 µm. (E) The frequency of two-celled proembryos with TUNEL-positive basal cells in WT and RNAi lines. Data represent the mean ± SE from three independent experiments, with 30 proembryos per line analysed in each experiment ( n = 90). (F–K) Morphology of the apical (G, J) and basal (H, K) cells analysed by TEM in the two-celled proembryos from WT (F–H) and RNAi lines (I–K). Scale bars, 5 µm in (F) and (I), and 2 µm in (G, H, J and K). Ac, apical cell; Bc, basal cell; n, nucleus; v, vacuole; nm, nuclear membrane; cw, cell wall; pm, plasma membrane. (L) Seed abortion in RNAi lines. Asterisks indicate aborted seeds. Scale bars, 1 mm. (M) Aberrant cell division patterns in the apical cell lineage at the early stages of embryogenesis in RNAi lines. Scale bars, 10 µm. (N) The frequency of aborted seeds in WT and RNAi lines. Data represent the mean ± SE from three independent experiments, with 400 to 500 seeds per line analysed in each experiment. (O, P) Cathepsin-like (O) and caspase-like (P) activities in the two-celled proembryos from WT and NtCYS RNAi line L2-6. Data represent the mean ± SE, with 40 to 60 two-celled proembryos. ** indicates statistical difference compared to WT ( t -test, p

    Journal: PLoS Biology

    Article Title: A Bipartite Molecular Module Controls Cell Death Activation in the Basal Cell Lineage of Plant EmbryosThe Fate of the Plant Embryo's Suspensor: Balancing Life and Death

    doi: 10.1371/journal.pbio.1001655

    Figure Lengend Snippet: Downregulation of NtCYS induces precocious cell death in the basal cell lineage and seed abortion. (A) Reduced expression of NtCYS in RNAi lines, as measured by RT-qPCR. The expression level of NtCYS in the WT was set to l. (B) The frequency of the two-celled proembryos with PI-positive basal cells in WT and RNAi lines. Data represent the mean ± SE from five independent experiments, with 50 proembryos per line analysed in each experiment ( n = 250). (C) Cell viability in the two-celled proembryos from WT and RNAi lines stained with FDA and PI. Scale bars, 10 µm. (D) Nuclear DNA fragmentation in the two-celled proembryos from WT and RNAi lines stained with TUNEL. Scale bars, 10 µm. (E) The frequency of two-celled proembryos with TUNEL-positive basal cells in WT and RNAi lines. Data represent the mean ± SE from three independent experiments, with 30 proembryos per line analysed in each experiment ( n = 90). (F–K) Morphology of the apical (G, J) and basal (H, K) cells analysed by TEM in the two-celled proembryos from WT (F–H) and RNAi lines (I–K). Scale bars, 5 µm in (F) and (I), and 2 µm in (G, H, J and K). Ac, apical cell; Bc, basal cell; n, nucleus; v, vacuole; nm, nuclear membrane; cw, cell wall; pm, plasma membrane. (L) Seed abortion in RNAi lines. Asterisks indicate aborted seeds. Scale bars, 1 mm. (M) Aberrant cell division patterns in the apical cell lineage at the early stages of embryogenesis in RNAi lines. Scale bars, 10 µm. (N) The frequency of aborted seeds in WT and RNAi lines. Data represent the mean ± SE from three independent experiments, with 400 to 500 seeds per line analysed in each experiment. (O, P) Cathepsin-like (O) and caspase-like (P) activities in the two-celled proembryos from WT and NtCYS RNAi line L2-6. Data represent the mean ± SE, with 40 to 60 two-celled proembryos. ** indicates statistical difference compared to WT ( t -test, p

    Article Snippet: The NtCYS and NtCP14 promoters were isolated using the Genome Walker DNA walking method with the Genome Walker Universal Kit (Clontech).

    Techniques: Expressing, Quantitative RT-PCR, Staining, TUNEL Assay, Transmission Electron Microscopy

    Constitutive overexpression of NtCP14 leads to cell death at the two-celled proembryo stage. (A) Enhanced expression of NtCP14 under the control of proZC1 promoter in transgenic lines, as measured by RT-qPCR. The expression level of NtCP14 in the WT was set to l. (B) Cathepsin H-like proteolytic activity towards substrate Bz-FVR-AMC in the two-celled proembryos from WT and NtCP14 -overexpressing line L14-4. Data represent the mean ± SE, with 45 to 65 two-celled proembryos in WT and line L14-4, respectively. (C) The frequency of aborted seeds in WT and NtCP14 -overexpressing lines. Data represent the mean ± SE from three independent experiments, with 300 to 400 seeds per line analysed in each experiment. (D) Cell viability in the two-celled proembryos from WT and NtCP14 -overexpressing line analysed by staining with FDA and PI. Scale bars, 10 µm. (E) The frequency of two-celled proembryos with PI-positive apical and basal cells in WT and NtCP14 -overexpressing lines. Data represent the mean ± SE from four independent experiments, with 50 proembryos per line analysed in each experiment ( n = 200). (F) Nuclear DNA fragmentation in the two-celled proembryos from WT and NtCP14 -overexpressing line L14-4 stained with TUNEL. Scale bars, 10 µm. (G) The frequency of two-celled proembryos with TUNEL-positive apical and basal cells in WT and NtCP14 -overexpressing line L14-4. Data represent the mean ± SE from three independent experiments, with 30 proembryos per line analysed in each experiment ( n = 90). (H) Caspase-like activities in the two-celled proembryos from WT and NtCP14 -overexpressing line L14-4. Data represent the mean ± SE, with 40 to 65 two-celled proembryos in WT and L14-4, respectively. (I) Cathepsin H-like activity of NtCP14 in the WT embryo proper versus suspensor at developmental stages 3 and 4. Data represent the mean ± SE, with 85 to 100 cells of embryo proper or suspensor analysed at stages 3 and 4. (J) Cathepsin H-like activity of NtCP14 in the basal cell lineage at successive stages of embryogenesis in WT plants. Data represent the mean ± SE, with 85 to 100 suspensor cells. * and ** indicate statistical difference compared to WT or as shown otherwise ( t -test, p

    Journal: PLoS Biology

    Article Title: A Bipartite Molecular Module Controls Cell Death Activation in the Basal Cell Lineage of Plant EmbryosThe Fate of the Plant Embryo's Suspensor: Balancing Life and Death

    doi: 10.1371/journal.pbio.1001655

    Figure Lengend Snippet: Constitutive overexpression of NtCP14 leads to cell death at the two-celled proembryo stage. (A) Enhanced expression of NtCP14 under the control of proZC1 promoter in transgenic lines, as measured by RT-qPCR. The expression level of NtCP14 in the WT was set to l. (B) Cathepsin H-like proteolytic activity towards substrate Bz-FVR-AMC in the two-celled proembryos from WT and NtCP14 -overexpressing line L14-4. Data represent the mean ± SE, with 45 to 65 two-celled proembryos in WT and line L14-4, respectively. (C) The frequency of aborted seeds in WT and NtCP14 -overexpressing lines. Data represent the mean ± SE from three independent experiments, with 300 to 400 seeds per line analysed in each experiment. (D) Cell viability in the two-celled proembryos from WT and NtCP14 -overexpressing line analysed by staining with FDA and PI. Scale bars, 10 µm. (E) The frequency of two-celled proembryos with PI-positive apical and basal cells in WT and NtCP14 -overexpressing lines. Data represent the mean ± SE from four independent experiments, with 50 proembryos per line analysed in each experiment ( n = 200). (F) Nuclear DNA fragmentation in the two-celled proembryos from WT and NtCP14 -overexpressing line L14-4 stained with TUNEL. Scale bars, 10 µm. (G) The frequency of two-celled proembryos with TUNEL-positive apical and basal cells in WT and NtCP14 -overexpressing line L14-4. Data represent the mean ± SE from three independent experiments, with 30 proembryos per line analysed in each experiment ( n = 90). (H) Caspase-like activities in the two-celled proembryos from WT and NtCP14 -overexpressing line L14-4. Data represent the mean ± SE, with 40 to 65 two-celled proembryos in WT and L14-4, respectively. (I) Cathepsin H-like activity of NtCP14 in the WT embryo proper versus suspensor at developmental stages 3 and 4. Data represent the mean ± SE, with 85 to 100 cells of embryo proper or suspensor analysed at stages 3 and 4. (J) Cathepsin H-like activity of NtCP14 in the basal cell lineage at successive stages of embryogenesis in WT plants. Data represent the mean ± SE, with 85 to 100 suspensor cells. * and ** indicate statistical difference compared to WT or as shown otherwise ( t -test, p

    Article Snippet: The NtCYS and NtCP14 promoters were isolated using the Genome Walker DNA walking method with the Genome Walker Universal Kit (Clontech).

    Techniques: Over Expression, Expressing, Transgenic Assay, Quantitative RT-PCR, Activity Assay, Staining, TUNEL Assay

    Effect of LCA treatment in HuH-7 cells on nuclear protein levels of Nrf2 and Mafs and ARE nuclear binding activity by these proteins. Part A ) Western blot analysis. Nuclear proteins were isolated from HuH-7 cells treated with LCA (100 µM) for up to 24 hours (15 µg protein/lane). Membranes were stripped and re-probed with histone 3 to ensure equal loading. Representative blots from 3 separate experiments are shown and numbers below the blots represent densitometric values expressed as % of 0 time control. Part B ) EMSA and supershift analysis. HuH-7 cells were treated with LCA (100 µM) or vehicle control for 20 hours and antibodies to Nrf1, Nrf2, MafG and c-Maf were used to compare relative amount of binding of these transcription factors to ARE as described in Methods. Part C ) ChIP analysis. HuH-7 cells were treated with LCA (100 µM) or vehicle control for 20 hours, then processed for ChIP assay as described in Methods. PCR products from amplification of the ARE sites following immunoprecipitation with antisera against Nrf2, Nrf1, c-Maf, MafG and Histone 3 demonstrate LCA treatment led to decreased Nrf2 and Nrf1 binding to the ARE sites, while c-Maf and MafG binding increased. Input genomic DNA (gDNA input) was used as a positive control and a no antibody immunoprecipitation (no Ab) was used as a negative control.

    Journal: Hepatology (Baltimore, Md.)

    Article Title: Induction of Maf Proteins by Toxic Bile Acid Inhibits Expression of GSH Synthetic Enzymes and Contributes to Cholestatic Liver Injury in Mice

    doi: 10.1002/hep.23471

    Figure Lengend Snippet: Effect of LCA treatment in HuH-7 cells on nuclear protein levels of Nrf2 and Mafs and ARE nuclear binding activity by these proteins. Part A ) Western blot analysis. Nuclear proteins were isolated from HuH-7 cells treated with LCA (100 µM) for up to 24 hours (15 µg protein/lane). Membranes were stripped and re-probed with histone 3 to ensure equal loading. Representative blots from 3 separate experiments are shown and numbers below the blots represent densitometric values expressed as % of 0 time control. Part B ) EMSA and supershift analysis. HuH-7 cells were treated with LCA (100 µM) or vehicle control for 20 hours and antibodies to Nrf1, Nrf2, MafG and c-Maf were used to compare relative amount of binding of these transcription factors to ARE as described in Methods. Part C ) ChIP analysis. HuH-7 cells were treated with LCA (100 µM) or vehicle control for 20 hours, then processed for ChIP assay as described in Methods. PCR products from amplification of the ARE sites following immunoprecipitation with antisera against Nrf2, Nrf1, c-Maf, MafG and Histone 3 demonstrate LCA treatment led to decreased Nrf2 and Nrf1 binding to the ARE sites, while c-Maf and MafG binding increased. Input genomic DNA (gDNA input) was used as a positive control and a no antibody immunoprecipitation (no Ab) was used as a negative control.

    Article Snippet: To clone the promoter region of the human GCLC gene, a reverse primer (5’-CAGCCTAATCTGGGAAATGAAGTTATTC -3’) corresponding to +427 to +455 (numbered according to the translational start site) of the human GCLC cDNA (GenBank® accession no. ) and a forward primer (5’-AGCAGCAGCAGCCCAGAGGTCAG -3’) corresponding to −3802 to −3779 (numbered according to the ATG start site) of our reported human GCLC 5’-flank region (GenBank® accession no. ) was used for PCR amplification (Advantage 2 PCR Enzyme system; BD Biosciences Clontech, Palo Alto, CA, U.S.A.) of a human genomic DNA (Genome Walker, Clontech).

    Techniques: Binding Assay, Activity Assay, Western Blot, Isolation, Chromatin Immunoprecipitation, Polymerase Chain Reaction, Amplification, Immunoprecipitation, Positive Control, Negative Control

    The T DNA insert is located in the promoter region of PIF3 . The diagram depicts (from top to bottom) the position of the poc1 locus on chromosome 1; the sequenced BAC F14J9 containing this locus; the T DNA insertion site in the promoter region of the PIF3 gene of poc1 and its location with respect to the predicted neighboring genes CAD and a MYB -related gene ( MYB ); the structure of the PIF3 gene and the location of the T DNA insert; and the structure of the PIF3 protein. The protein coding regions within the PIF3 gene are shown as filled boxes, the untranslated regions as open boxes, the introns and flanking DNA as a line, and the putative TATA box as vertical lines. The identity and location of motifs within the PIF3 protein are shown: PAS, Per–Arnt–Sim-like domain; bHLH, basic helix–loop–helix domain; NLS, nuclear localization signal. The position of the probe used in quantitative PCR analysis of PIF3 transcript levels also is shown (Probe).

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: poc1: An Arabidopsis mutant perturbed in phytochrome signaling because of a T DNA insertion in the promoter of PIF3, a gene encoding a phytochrome-interacting bHLH protein

    doi:

    Figure Lengend Snippet: The T DNA insert is located in the promoter region of PIF3 . The diagram depicts (from top to bottom) the position of the poc1 locus on chromosome 1; the sequenced BAC F14J9 containing this locus; the T DNA insertion site in the promoter region of the PIF3 gene of poc1 and its location with respect to the predicted neighboring genes CAD and a MYB -related gene ( MYB ); the structure of the PIF3 gene and the location of the T DNA insert; and the structure of the PIF3 protein. The protein coding regions within the PIF3 gene are shown as filled boxes, the untranslated regions as open boxes, the introns and flanking DNA as a line, and the putative TATA box as vertical lines. The identity and location of motifs within the PIF3 protein are shown: PAS, Per–Arnt–Sim-like domain; bHLH, basic helix–loop–helix domain; NLS, nuclear localization signal. The position of the probe used in quantitative PCR analysis of PIF3 transcript levels also is shown (Probe).

    Article Snippet: Genomic DNA flanking the right border was cloned by using the PCR-based Genome Walker kit (CLONTECH) and sequenced.

    Techniques: BAC Assay, Real-time Polymerase Chain Reaction