activation domain fusion vectors pgbkt7 Takara Search Results


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  • 85
    TaKaRa gal4 dna binding domain fusions
    Interaction of the Nac1 and Miz1 POZ domains in yeast two-hybrid assays AH109 yeast cells were transformed with constructs expressing <t>GAL4</t> activation domain fusion proteins (pGADT7) together with GAL4 <t>DNA-binding</t> domain fusion proteins (pGBKT7) and plated onto media lacking leucine and tryptophan [-LT], lacking leucine, tryptophan and histidine [-HLT], and lacking leucine, tryptophan, histidine and adenine [-LTHA]. Transformation with Miz1POZ-pGADT7 together with: (1) pGBKT7, (2) Zbtb8 POZ-pGBKT7, (3) Zbtb6 POZ-pGBKT7, (6) Nac1 POZ-pGBKT7, and (7) BCL6 POZ-pGBKT7 (7). Transformation with Nac1 POZ-pGBKT7 together with: (4) Zbtb8 POZ-pGADT7, (5) Zbtb6-pGADT7 and (8) pGADT7.
    Gal4 Dna Binding Domain Fusions, supplied by TaKaRa, used in various techniques. Bioz Stars score: 85/100, based on 177 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    TaKaRa pgbkt7
    FANCC directly interacts with UNC5A in yeasts. Yeast two-hybrid assays were performed with various UNC5A constructs that included the Y2H clone coding for the C-terminal death domain (DEATH) and a part of intron 14 (shown in blue), UNC5A intracellular domain (UNC5A ICD ), the UNC5A C-terminus (UNC5A DD ) and the death-domain deletion mutant (UNC5A ΔDD ). The yeast strain AH109 was co-transformed with UNC5A constructs along with FANCC constructs as indicated. The plus sign (+) indicate a positive interaction. Negative controls were performed using empty vectors (Empty), and positive controls were performed using <t>pGBKT7/p53</t> and pGADT7/SV40 T-antigen from Clontech (data not shown). Yeast two-hybrid assays were performed at least three times (independent transformation) with 3 to 4 clones each.
    Pgbkt7, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 6016 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    TaKaRa pgbkt7 vector
    Effect of mutation p.Ala449Thr on the GK interaction with GKRP. A) Inhibition of glucokinase activity by human GKRP. Enzyme activity was measured at 5 mM glucose as described in Material and Methods , in the presence of 10 µM S6P (left panel) or 0.2 mM F1P (right panel). Results are means ± SEM for three independent enzyme purifications assayed in triplicate. B) Two-hybrid interaction of GBD-GKRP with GAD-GK or GAD-GK(p.Ala449Thr) mutant. Yeast strain Y187 was used, and fusion proteins were expressed from <t>pGBKT7</t> and pACTII derivatives. Values are means ± SEM from ß-galactosidase activity of six independent transformants. In control experiments, GBD-GKRP did not interact with GAD and GAD-GK did not interact with GBD [22] .
    Pgbkt7 Vector, supplied by TaKaRa, used in various techniques. Bioz Stars score: 90/100, based on 2385 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    98
    TaKaRa pactii
    Effect of mutation p.Ala449Thr on the GK interaction with GKRP. A) Inhibition of glucokinase activity by human GKRP. Enzyme activity was measured at 5 mM glucose as described in Material and Methods , in the presence of 10 µM S6P (left panel) or 0.2 mM F1P (right panel). Results are means ± SEM for three independent enzyme purifications assayed in triplicate. B) Two-hybrid interaction of GBD-GKRP with <t>GAD-GK</t> or GAD-GK(p.Ala449Thr) mutant. Yeast strain Y187 was used, and fusion proteins were expressed from pGBKT7 and <t>pACTII</t> derivatives. Values are means ± SEM from ß-galactosidase activity of six independent transformants. In control experiments, GBD-GKRP did not interact with GAD and GAD-GK did not interact with GBD [22] .
    Pactii, supplied by TaKaRa, used in various techniques. Bioz Stars score: 98/100, based on 193 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TaKaRa gal4 activation domain
    Effect of mutation p.Ala449Thr on the GK interaction with GKRP. A) Inhibition of glucokinase activity by human GKRP. Enzyme activity was measured at 5 mM glucose as described in Material and Methods , in the presence of 10 µM S6P (left panel) or 0.2 mM F1P (right panel). Results are means ± SEM for three independent enzyme purifications assayed in triplicate. B) Two-hybrid interaction of GBD-GKRP with <t>GAD-GK</t> or GAD-GK(p.Ala449Thr) mutant. Yeast strain Y187 was used, and fusion proteins were expressed from pGBKT7 and <t>pACTII</t> derivatives. Values are means ± SEM from ß-galactosidase activity of six independent transformants. In control experiments, GBD-GKRP did not interact with GAD and GAD-GK did not interact with GBD [22] .
    Gal4 Activation Domain, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 1413 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    TaKaRa ah109
    Effect of mutation p.Ala449Thr on the GK interaction with GKRP. A) Inhibition of glucokinase activity by human GKRP. Enzyme activity was measured at 5 mM glucose as described in Material and Methods , in the presence of 10 µM S6P (left panel) or 0.2 mM F1P (right panel). Results are means ± SEM for three independent enzyme purifications assayed in triplicate. B) Two-hybrid interaction of GBD-GKRP with <t>GAD-GK</t> or GAD-GK(p.Ala449Thr) mutant. Yeast strain Y187 was used, and fusion proteins were expressed from pGBKT7 and <t>pACTII</t> derivatives. Values are means ± SEM from ß-galactosidase activity of six independent transformants. In control experiments, GBD-GKRP did not interact with GAD and GAD-GK did not interact with GBD [22] .
    Ah109, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 468 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    95
    TaKaRa pgbkt7 p53
    Effect of mutation p.Ala449Thr on the GK interaction with GKRP. A) Inhibition of glucokinase activity by human GKRP. Enzyme activity was measured at 5 mM glucose as described in Material and Methods , in the presence of 10 µM S6P (left panel) or 0.2 mM F1P (right panel). Results are means ± SEM for three independent enzyme purifications assayed in triplicate. B) Two-hybrid interaction of GBD-GKRP with <t>GAD-GK</t> or GAD-GK(p.Ala449Thr) mutant. Yeast strain Y187 was used, and fusion proteins were expressed from pGBKT7 and <t>pACTII</t> derivatives. Values are means ± SEM from ß-galactosidase activity of six independent transformants. In control experiments, GBD-GKRP did not interact with GAD and GAD-GK did not interact with GBD [22] .
    Pgbkt7 P53, supplied by TaKaRa, used in various techniques. Bioz Stars score: 95/100, based on 136 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    TaKaRa pgadt7 t
    Effect of mutation p.Ala449Thr on the GK interaction with GKRP. A) Inhibition of glucokinase activity by human GKRP. Enzyme activity was measured at 5 mM glucose as described in Material and Methods , in the presence of 10 µM S6P (left panel) or 0.2 mM F1P (right panel). Results are means ± SEM for three independent enzyme purifications assayed in triplicate. B) Two-hybrid interaction of GBD-GKRP with <t>GAD-GK</t> or GAD-GK(p.Ala449Thr) mutant. Yeast strain Y187 was used, and fusion proteins were expressed from pGBKT7 and <t>pACTII</t> derivatives. Values are means ± SEM from ß-galactosidase activity of six independent transformants. In control experiments, GBD-GKRP did not interact with GAD and GAD-GK did not interact with GBD [22] .
    Pgadt7 T, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 349 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TaKaRa matchmaker gal4 two hybrid system 3
    Effect of mutation p.Ala449Thr on the GK interaction with GKRP. A) Inhibition of glucokinase activity by human GKRP. Enzyme activity was measured at 5 mM glucose as described in Material and Methods , in the presence of 10 µM S6P (left panel) or 0.2 mM F1P (right panel). Results are means ± SEM for three independent enzyme purifications assayed in triplicate. B) Two-hybrid interaction of GBD-GKRP with <t>GAD-GK</t> or GAD-GK(p.Ala449Thr) mutant. Yeast strain Y187 was used, and fusion proteins were expressed from pGBKT7 and <t>pACTII</t> derivatives. Values are means ± SEM from ß-galactosidase activity of six independent transformants. In control experiments, GBD-GKRP did not interact with GAD and GAD-GK did not interact with GBD [22] .
    Matchmaker Gal4 Two Hybrid System 3, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 1141 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    97
    TaKaRa strain ah109
    Effect of mutation p.Ala449Thr on the GK interaction with GKRP. A) Inhibition of glucokinase activity by human GKRP. Enzyme activity was measured at 5 mM glucose as described in Material and Methods , in the presence of 10 µM S6P (left panel) or 0.2 mM F1P (right panel). Results are means ± SEM for three independent enzyme purifications assayed in triplicate. B) Two-hybrid interaction of GBD-GKRP with <t>GAD-GK</t> or GAD-GK(p.Ala449Thr) mutant. Yeast strain Y187 was used, and fusion proteins were expressed from pGBKT7 and <t>pACTII</t> derivatives. Values are means ± SEM from ß-galactosidase activity of six independent transformants. In control experiments, GBD-GKRP did not interact with GAD and GAD-GK did not interact with GBD [22] .
    Strain Ah109, supplied by TaKaRa, used in various techniques. Bioz Stars score: 97/100, based on 135 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TaKaRa human hacat cdna library
    Effect of mutation p.Ala449Thr on the GK interaction with GKRP. A) Inhibition of glucokinase activity by human GKRP. Enzyme activity was measured at 5 mM glucose as described in Material and Methods , in the presence of 10 µM S6P (left panel) or 0.2 mM F1P (right panel). Results are means ± SEM for three independent enzyme purifications assayed in triplicate. B) Two-hybrid interaction of GBD-GKRP with <t>GAD-GK</t> or GAD-GK(p.Ala449Thr) mutant. Yeast strain Y187 was used, and fusion proteins were expressed from pGBKT7 and <t>pACTII</t> derivatives. Values are means ± SEM from ß-galactosidase activity of six independent transformants. In control experiments, GBD-GKRP did not interact with GAD and GAD-GK did not interact with GBD [22] .
    Human Hacat Cdna Library, supplied by TaKaRa, used in various techniques. Bioz Stars score: 78/100, based on 6 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    gal4  (TaKaRa)
    90
    TaKaRa gal4
    E2E1, E2G1, E2J1, E2J2, and E2L3 interact with MuRF1 in the presence of telethonin (A) Telethonin does not interact with E2s. Y2H experiments were performed using telethonin as a bait to confirm that this protein cannot directly interact with the E2 enzymes used in this work. The empty vector and the vector containing the LT construct were used as negative controls against telethonin to estimate potential background level. Signals above ‘empty’ and ‘LT’ lanes were considered as positive. Colonies were plated on selective medium [ ‐LTH + Aureo + 3‐AT ] (Experimental section) and monitored during 21 days. LT, Large‐T antigen; Tele, telethonin. (B) Telethonin expression level in yeast varies a ccording to methionine (Met) concentration in the medium. BD‐Tele, fusion protein between the binding domain of <t>GAL4</t> and telethonin; Tele, telethonin. (C) Densitometry analysis from the immunoblot presented in (B). (D) Yeast three‐hybrid (Y3H) experiments revealed E2E1, E2G1, E2J1, E2J2, and E2L3 as MuRF1 partners in the presence of telethonin. E2‐expressing yeasts were mated against strains expressing either MuRF1 alone or MuRF1 and telethonin. Colonies were plated on selective medium [ ‐LTH + Aureo + 3‐AT ] containing 134 mM Met. Results were observed at day 6. Three to four independent transformation experiments were performed and 11 to 32 colonies were analyzed for each E2.
    Gal4, supplied by TaKaRa, used in various techniques. Bioz Stars score: 90/100, based on 187 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TaKaRa pact2 vector
    N protein of SARS-CoV associates with EF1α. (A) Interaction of N protein with EF1α was revealed by a yeast two-hybrid system. Three transformants containing pGBKT7-N and <t>pACT2-EF1α(291-463)</t> (left panel) together with both positive (upper right) and negative (lower right) controls were inoculated on SD-Leu − Trp − His − Ade − -X-α-Gal plates. Transformants containing pGADT7-T and pGBKT7-p53 were used as a positive control, and transformants containing pGADT7-T and a pGBKT7-Lam were employed as a negative control. (B) Lysates from 293T cells transfected with Flag-EF1α or Flag and GFP-N or GFP plasmids were subjected to immunoprecipitation with anti-Flag antibody and analyzed by immunoblotting with anti-GFP or anti-Flag antibody. (C) Lysates from 293T cells transfected with Flag-N or Flag plasmid were subjected to immunoprecipitation with anti-Flag and analyzed by immunoblotting with anti-EF1α antibody. (D) Lysates from 293T cells transfected with GFP-N or the indicated GFP-N truncation forms and Flag-EF1α were subjected to immunoprecipitation with anti-Flag and subsequently analyzed by immunoblotting with anti-GFP or anti-Flag antibody. IP, immunoprecipitation; IB, immunoblotting; P/N, positive/negative.
    Pact2 Vector, supplied by TaKaRa, used in various techniques. Bioz Stars score: 99/100, based on 833 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TaKaRa gal4 dbd
    N protein of SARS-CoV associates with EF1α. (A) Interaction of N protein with EF1α was revealed by a yeast two-hybrid system. Three transformants containing pGBKT7-N and <t>pACT2-EF1α(291-463)</t> (left panel) together with both positive (upper right) and negative (lower right) controls were inoculated on SD-Leu − Trp − His − Ade − -X-α-Gal plates. Transformants containing pGADT7-T and pGBKT7-p53 were used as a positive control, and transformants containing pGADT7-T and a pGBKT7-Lam were employed as a negative control. (B) Lysates from 293T cells transfected with Flag-EF1α or Flag and GFP-N or GFP plasmids were subjected to immunoprecipitation with anti-Flag antibody and analyzed by immunoblotting with anti-GFP or anti-Flag antibody. (C) Lysates from 293T cells transfected with Flag-N or Flag plasmid were subjected to immunoprecipitation with anti-Flag and analyzed by immunoblotting with anti-EF1α antibody. (D) Lysates from 293T cells transfected with GFP-N or the indicated GFP-N truncation forms and Flag-EF1α were subjected to immunoprecipitation with anti-Flag and subsequently analyzed by immunoblotting with anti-GFP or anti-Flag antibody. IP, immunoprecipitation; IB, immunoblotting; P/N, positive/negative.
    Gal4 Dbd, supplied by TaKaRa, used in various techniques. Bioz Stars score: 98/100, based on 220 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TaKaRa matchmaker gold yeast two hybrid system
    N protein of SARS-CoV associates with EF1α. (A) Interaction of N protein with EF1α was revealed by a yeast two-hybrid system. Three transformants containing pGBKT7-N and <t>pACT2-EF1α(291-463)</t> (left panel) together with both positive (upper right) and negative (lower right) controls were inoculated on SD-Leu − Trp − His − Ade − -X-α-Gal plates. Transformants containing pGADT7-T and pGBKT7-p53 were used as a positive control, and transformants containing pGADT7-T and a pGBKT7-Lam were employed as a negative control. (B) Lysates from 293T cells transfected with Flag-EF1α or Flag and GFP-N or GFP plasmids were subjected to immunoprecipitation with anti-Flag antibody and analyzed by immunoblotting with anti-GFP or anti-Flag antibody. (C) Lysates from 293T cells transfected with Flag-N or Flag plasmid were subjected to immunoprecipitation with anti-Flag and analyzed by immunoblotting with anti-EF1α antibody. (D) Lysates from 293T cells transfected with GFP-N or the indicated GFP-N truncation forms and Flag-EF1α were subjected to immunoprecipitation with anti-Flag and subsequently analyzed by immunoblotting with anti-GFP or anti-Flag antibody. IP, immunoprecipitation; IB, immunoblotting; P/N, positive/negative.
    Matchmaker Gold Yeast Two Hybrid System, supplied by TaKaRa, used in various techniques. Bioz Stars score: 90/100, based on 1551 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TaKaRa ecori
    N protein of SARS-CoV associates with EF1α. (A) Interaction of N protein with EF1α was revealed by a yeast two-hybrid system. Three transformants containing pGBKT7-N and <t>pACT2-EF1α(291-463)</t> (left panel) together with both positive (upper right) and negative (lower right) controls were inoculated on SD-Leu − Trp − His − Ade − -X-α-Gal plates. Transformants containing pGADT7-T and pGBKT7-p53 were used as a positive control, and transformants containing pGADT7-T and a pGBKT7-Lam were employed as a negative control. (B) Lysates from 293T cells transfected with Flag-EF1α or Flag and GFP-N or GFP plasmids were subjected to immunoprecipitation with anti-Flag antibody and analyzed by immunoblotting with anti-GFP or anti-Flag antibody. (C) Lysates from 293T cells transfected with Flag-N or Flag plasmid were subjected to immunoprecipitation with anti-Flag and analyzed by immunoblotting with anti-EF1α antibody. (D) Lysates from 293T cells transfected with GFP-N or the indicated GFP-N truncation forms and Flag-EF1α were subjected to immunoprecipitation with anti-Flag and subsequently analyzed by immunoblotting with anti-GFP or anti-Flag antibody. IP, immunoprecipitation; IB, immunoblotting; P/N, positive/negative.
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    TaKaRa library screen yeast two hybrid assay
    N protein of SARS-CoV associates with EF1α. (A) Interaction of N protein with EF1α was revealed by a yeast two-hybrid system. Three transformants containing pGBKT7-N and <t>pACT2-EF1α(291-463)</t> (left panel) together with both positive (upper right) and negative (lower right) controls were inoculated on SD-Leu − Trp − His − Ade − -X-α-Gal plates. Transformants containing pGADT7-T and pGBKT7-p53 were used as a positive control, and transformants containing pGADT7-T and a pGBKT7-Lam were employed as a negative control. (B) Lysates from 293T cells transfected with Flag-EF1α or Flag and GFP-N or GFP plasmids were subjected to immunoprecipitation with anti-Flag antibody and analyzed by immunoblotting with anti-GFP or anti-Flag antibody. (C) Lysates from 293T cells transfected with Flag-N or Flag plasmid were subjected to immunoprecipitation with anti-Flag and analyzed by immunoblotting with anti-EF1α antibody. (D) Lysates from 293T cells transfected with GFP-N or the indicated GFP-N truncation forms and Flag-EF1α were subjected to immunoprecipitation with anti-Flag and subsequently analyzed by immunoblotting with anti-GFP or anti-Flag antibody. IP, immunoprecipitation; IB, immunoblotting; P/N, positive/negative.
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    81
    TaKaRa pgadt7 yeast vectors
    FANCC directly interacts with UNC5A in yeasts. Yeast two-hybrid assays were performed with various UNC5A constructs that included the Y2H clone coding for the C-terminal death domain (DEATH) and a part of intron 14 (shown in blue), UNC5A intracellular domain (UNC5A ICD ), the UNC5A C-terminus (UNC5A DD ) and the death-domain deletion mutant (UNC5A ΔDD ). The yeast strain AH109 was co-transformed with UNC5A constructs along with FANCC constructs as indicated. The plus sign (+) indicate a positive interaction. Negative controls were performed using empty vectors (Empty), and positive controls were performed using pGBKT7/p53 and <t>pGADT7/SV40</t> T-antigen from Clontech (data not shown). Yeast two-hybrid assays were performed at least three times (independent transformation) with 3 to 4 clones each.
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    90
    TaKaRa human bone marrow cdna library
    FANCC directly interacts with UNC5A in yeasts. Yeast two-hybrid assays were performed with various UNC5A constructs that included the Y2H clone coding for the C-terminal death domain (DEATH) and a part of intron 14 (shown in blue), UNC5A intracellular domain (UNC5A ICD ), the UNC5A C-terminus (UNC5A DD ) and the death-domain deletion mutant (UNC5A ΔDD ). The yeast strain AH109 was co-transformed with UNC5A constructs along with FANCC constructs as indicated. The plus sign (+) indicate a positive interaction. Negative controls were performed using empty vectors (Empty), and positive controls were performed using pGBKT7/p53 and <t>pGADT7/SV40</t> T-antigen from Clontech (data not shown). Yeast two-hybrid assays were performed at least three times (independent transformation) with 3 to 4 clones each.
    Human Bone Marrow Cdna Library, supplied by TaKaRa, used in various techniques. Bioz Stars score: 90/100, based on 21 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    90
    TaKaRa bamhi
    FANCC directly interacts with UNC5A in yeasts. Yeast two-hybrid assays were performed with various UNC5A constructs that included the Y2H clone coding for the C-terminal death domain (DEATH) and a part of intron 14 (shown in blue), UNC5A intracellular domain (UNC5A ICD ), the UNC5A C-terminus (UNC5A DD ) and the death-domain deletion mutant (UNC5A ΔDD ). The yeast strain AH109 was co-transformed with UNC5A constructs along with FANCC constructs as indicated. The plus sign (+) indicate a positive interaction. Negative controls were performed using empty vectors (Empty), and positive controls were performed using pGBKT7/p53 and <t>pGADT7/SV40</t> T-antigen from Clontech (data not shown). Yeast two-hybrid assays were performed at least three times (independent transformation) with 3 to 4 clones each.
    Bamhi, supplied by TaKaRa, used in various techniques. Bioz Stars score: 90/100, based on 5292 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    TaKaRa dna constructs
    FANCC directly interacts with UNC5A in yeasts. Yeast two-hybrid assays were performed with various UNC5A constructs that included the Y2H clone coding for the C-terminal death domain (DEATH) and a part of intron 14 (shown in blue), UNC5A intracellular domain (UNC5A ICD ), the UNC5A C-terminus (UNC5A DD ) and the death-domain deletion mutant (UNC5A ΔDD ). The yeast strain AH109 was co-transformed with UNC5A constructs along with FANCC constructs as indicated. The plus sign (+) indicate a positive interaction. Negative controls were performed using empty vectors (Empty), and positive controls were performed using pGBKT7/p53 and <t>pGADT7/SV40</t> T-antigen from Clontech (data not shown). Yeast two-hybrid assays were performed at least three times (independent transformation) with 3 to 4 clones each.
    Dna Constructs, supplied by TaKaRa, used in various techniques. Bioz Stars score: 93/100, based on 699 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    TaKaRa yeast haploid strains y187
    FANCC directly interacts with UNC5A in yeasts. Yeast two-hybrid assays were performed with various UNC5A constructs that included the Y2H clone coding for the C-terminal death domain (DEATH) and a part of intron 14 (shown in blue), UNC5A intracellular domain (UNC5A ICD ), the UNC5A C-terminus (UNC5A DD ) and the death-domain deletion mutant (UNC5A ΔDD ). The yeast strain AH109 was co-transformed with UNC5A constructs along with FANCC constructs as indicated. The plus sign (+) indicate a positive interaction. Negative controls were performed using empty vectors (Empty), and positive controls were performed using pGBKT7/p53 and <t>pGADT7/SV40</t> T-antigen from Clontech (data not shown). Yeast two-hybrid assays were performed at least three times (independent transformation) with 3 to 4 clones each.
    Yeast Haploid Strains Y187, supplied by TaKaRa, used in various techniques. Bioz Stars score: 90/100, based on 14 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/yeast haploid strains y187/product/TaKaRa
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    Interaction of the Nac1 and Miz1 POZ domains in yeast two-hybrid assays AH109 yeast cells were transformed with constructs expressing GAL4 activation domain fusion proteins (pGADT7) together with GAL4 DNA-binding domain fusion proteins (pGBKT7) and plated onto media lacking leucine and tryptophan [-LT], lacking leucine, tryptophan and histidine [-HLT], and lacking leucine, tryptophan, histidine and adenine [-LTHA]. Transformation with Miz1POZ-pGADT7 together with: (1) pGBKT7, (2) Zbtb8 POZ-pGBKT7, (3) Zbtb6 POZ-pGBKT7, (6) Nac1 POZ-pGBKT7, and (7) BCL6 POZ-pGBKT7 (7). Transformation with Nac1 POZ-pGBKT7 together with: (4) Zbtb8 POZ-pGADT7, (5) Zbtb6-pGADT7 and (8) pGADT7.

    Journal: Bioscience Reports

    Article Title: Nac1 interacts with the POZ-domain transcription factor, Miz1

    doi: 10.1042/BSR20140049

    Figure Lengend Snippet: Interaction of the Nac1 and Miz1 POZ domains in yeast two-hybrid assays AH109 yeast cells were transformed with constructs expressing GAL4 activation domain fusion proteins (pGADT7) together with GAL4 DNA-binding domain fusion proteins (pGBKT7) and plated onto media lacking leucine and tryptophan [-LT], lacking leucine, tryptophan and histidine [-HLT], and lacking leucine, tryptophan, histidine and adenine [-LTHA]. Transformation with Miz1POZ-pGADT7 together with: (1) pGBKT7, (2) Zbtb8 POZ-pGBKT7, (3) Zbtb6 POZ-pGBKT7, (6) Nac1 POZ-pGBKT7, and (7) BCL6 POZ-pGBKT7 (7). Transformation with Nac1 POZ-pGBKT7 together with: (4) Zbtb8 POZ-pGADT7, (5) Zbtb6-pGADT7 and (8) pGADT7.

    Article Snippet: The mouse Miz1 POZ domain (residues 1–115) was expressed as a GAL4 activation domain fusion using the vector pGADT7 (Clontech Laboratories Inc.), and the POZ domains of 32 mouse transcription factors were expressed as GAL4 DNA-binding domain fusions using pGBKT7 (Clontech Laboratories Inc.).

    Techniques: Transformation Assay, Construct, Expressing, Activation Assay, Binding Assay

    FANCC directly interacts with UNC5A in yeasts. Yeast two-hybrid assays were performed with various UNC5A constructs that included the Y2H clone coding for the C-terminal death domain (DEATH) and a part of intron 14 (shown in blue), UNC5A intracellular domain (UNC5A ICD ), the UNC5A C-terminus (UNC5A DD ) and the death-domain deletion mutant (UNC5A ΔDD ). The yeast strain AH109 was co-transformed with UNC5A constructs along with FANCC constructs as indicated. The plus sign (+) indicate a positive interaction. Negative controls were performed using empty vectors (Empty), and positive controls were performed using pGBKT7/p53 and pGADT7/SV40 T-antigen from Clontech (data not shown). Yeast two-hybrid assays were performed at least three times (independent transformation) with 3 to 4 clones each.

    Journal: PLoS ONE

    Article Title: The Fanconi Anemia Group C Protein Interacts with Uncoordinated 5A and Delays Apoptosis

    doi: 10.1371/journal.pone.0092811

    Figure Lengend Snippet: FANCC directly interacts with UNC5A in yeasts. Yeast two-hybrid assays were performed with various UNC5A constructs that included the Y2H clone coding for the C-terminal death domain (DEATH) and a part of intron 14 (shown in blue), UNC5A intracellular domain (UNC5A ICD ), the UNC5A C-terminus (UNC5A DD ) and the death-domain deletion mutant (UNC5A ΔDD ). The yeast strain AH109 was co-transformed with UNC5A constructs along with FANCC constructs as indicated. The plus sign (+) indicate a positive interaction. Negative controls were performed using empty vectors (Empty), and positive controls were performed using pGBKT7/p53 and pGADT7/SV40 T-antigen from Clontech (data not shown). Yeast two-hybrid assays were performed at least three times (independent transformation) with 3 to 4 clones each.

    Article Snippet: Plasmids and DNA constructs The N-terminus of FANCC, which spans from nucleotides 256 to 1175 and encompasses amino acids from the start codon to the cleavage site , was cloned into the pGBKT7 and pGADT7 yeast vectors (Clontech Laboratories Inc., Mountain View, CA) by fusion to the Gal4-DNA binding or DNA-activating domain, and into the pEGFP plasmids (pGBKFANCC1-306 , pGADFANCC1-306 , pEGFPFANCC1-306 ).

    Techniques: Construct, Mutagenesis, Transformation Assay, Clone Assay

    Autoactivation of three reporter genes by MVLG_04106 on QDO/X-α-gal + 3-AT (5 mM) plates. Undil, undiluted; 10× and 100× dilutions. QDO (Quadruple drop out media), 3-AT (3-Amino-1,2,4-triazole), BD (DNA binding domain in pGBKT7 vector), AD (Activation domain in pGADT7 vector), BD-p53 (pGBKT7-53 positive control plasmid), AD-T (pGADT7-T positive control plasmid), and BD-4106∆SP (MVLG_04106 lacking signal peptide).

    Journal: International Journal of Molecular Sciences

    Article Title: Identification and Initial Characterization of the Effectors of an Anther Smut Fungus and Potential Host Target Proteins

    doi: 10.3390/ijms18112489

    Figure Lengend Snippet: Autoactivation of three reporter genes by MVLG_04106 on QDO/X-α-gal + 3-AT (5 mM) plates. Undil, undiluted; 10× and 100× dilutions. QDO (Quadruple drop out media), 3-AT (3-Amino-1,2,4-triazole), BD (DNA binding domain in pGBKT7 vector), AD (Activation domain in pGADT7 vector), BD-p53 (pGBKT7-53 positive control plasmid), AD-T (pGADT7-T positive control plasmid), and BD-4106∆SP (MVLG_04106 lacking signal peptide).

    Article Snippet: Interactors with bait are identified by screening “prey” expressed from a yeast vector where the fusion is with the Gal4 transcriptional activation domain (AD). pGBKT7 was used as a “bait” vector with the GAL4 DNA-binding domain and pGADT7 was used as a “prey” vector with the GAL4 DNA activation domain.

    Techniques: Binding Assay, Plasmid Preparation, Activation Assay, Positive Control

    Identification of hDUS2 as PKR and PACT interacting protein. ( A ) Yeast two-hybrid interaction assay. The indicated plasmids were transformed into yeast strain Hf7C and the transformants were streaked on triple dropout medium lacking leucine, tryptophan and histidine. A: hDUS2/pGBKT7 and empty vector pGADT7, B: hDUS2/pGBKT7 and PKR/pGADT7, C: hDUS2/pGBKT7 and PACT/pGADT7, D: empty vector pGBKT7 and hDUS2/pGADT7, E: PKR/pGBKT7 and hDUS2/pGADT7, F: PACT/pGBKT7 and hDUS2/pGADT7. ( B ) β-Galactosidase assay for the interactions in yeast. The indicated plasmids were transformed into yeast strain Hf7C and the transformants were streaked on double dropout medium lacking leucine, and tryptophan. After 4 days the growth was lifted on nitrocellulose membrane and β-galactosidase activity assay was performed after lysis of yeast cells on the membrane. Blue color indicates a positive interaction and white color indicates no interaction. ( C ) Biochemical interaction assay. In vitro -translated hDUS2 protein was allowed to interact with hexahistidine-tagged pure recombinant PKR or PACT proteins that were bound to Ni-agarose affinity beads. The bound proteins remaining on the beads were analyzed by SDS–PAGE followed by phosphorimager analysis. Lane 1: total protein from the translation mix (20% of input in lanes 2–4), lanes 2–4: hDUS2 protein bound to beads. Lane 2: protein bound to his-DRIL1 beads: negative control, lane 3: protein bound to his-PKR beads, lane 4: protein bound to his-PACT beads. ( D ) Domain structure of hDUS2. A schematic representation of the DUS and dsRBM domains in hDUS2 protein. The DUS domain is shown as a hatched box and the dsRBM is shown as a gray box. The amino acid numbers are indicated on the top and bottom of the boxes. ( E ) Alignment of hDUS2 dsRBM with dsRBMs from three other human dsRNA-binding proteins PKR, PACT and TRBP. ClustalW alignment of the dsRBMs present in PKR, PACT and TRBP is shown. The conserved residues are shown in dark gray and similarities are shown in light gray. The consensus is shown at the bottom.

    Journal: Nucleic Acids Research

    Article Title: Interaction of human tRNA-dihydrouridine synthase-2 with interferon-induced protein kinase PKR

    doi: 10.1093/nar/gkm1129

    Figure Lengend Snippet: Identification of hDUS2 as PKR and PACT interacting protein. ( A ) Yeast two-hybrid interaction assay. The indicated plasmids were transformed into yeast strain Hf7C and the transformants were streaked on triple dropout medium lacking leucine, tryptophan and histidine. A: hDUS2/pGBKT7 and empty vector pGADT7, B: hDUS2/pGBKT7 and PKR/pGADT7, C: hDUS2/pGBKT7 and PACT/pGADT7, D: empty vector pGBKT7 and hDUS2/pGADT7, E: PKR/pGBKT7 and hDUS2/pGADT7, F: PACT/pGBKT7 and hDUS2/pGADT7. ( B ) β-Galactosidase assay for the interactions in yeast. The indicated plasmids were transformed into yeast strain Hf7C and the transformants were streaked on double dropout medium lacking leucine, and tryptophan. After 4 days the growth was lifted on nitrocellulose membrane and β-galactosidase activity assay was performed after lysis of yeast cells on the membrane. Blue color indicates a positive interaction and white color indicates no interaction. ( C ) Biochemical interaction assay. In vitro -translated hDUS2 protein was allowed to interact with hexahistidine-tagged pure recombinant PKR or PACT proteins that were bound to Ni-agarose affinity beads. The bound proteins remaining on the beads were analyzed by SDS–PAGE followed by phosphorimager analysis. Lane 1: total protein from the translation mix (20% of input in lanes 2–4), lanes 2–4: hDUS2 protein bound to beads. Lane 2: protein bound to his-DRIL1 beads: negative control, lane 3: protein bound to his-PKR beads, lane 4: protein bound to his-PACT beads. ( D ) Domain structure of hDUS2. A schematic representation of the DUS and dsRBM domains in hDUS2 protein. The DUS domain is shown as a hatched box and the dsRBM is shown as a gray box. The amino acid numbers are indicated on the top and bottom of the boxes. ( E ) Alignment of hDUS2 dsRBM with dsRBMs from three other human dsRNA-binding proteins PKR, PACT and TRBP. ClustalW alignment of the dsRBMs present in PKR, PACT and TRBP is shown. The conserved residues are shown in dark gray and similarities are shown in light gray. The consensus is shown at the bottom.

    Article Snippet: The full-length open reading frame for hDUS2 was sub-cloned into the yeast two hybrid vectors pGBKT7 and pGADT7 to obtain GAL4 DNA-binding domain fusion and GAL4 activation domain fusion, respectively.

    Techniques: Transformation Assay, Plasmid Preparation, Activity Assay, Lysis, In Vitro, Recombinant, SDS Page, Negative Control, Binding Assay

    Effect of mutation p.Ala449Thr on the GK interaction with GKRP. A) Inhibition of glucokinase activity by human GKRP. Enzyme activity was measured at 5 mM glucose as described in Material and Methods , in the presence of 10 µM S6P (left panel) or 0.2 mM F1P (right panel). Results are means ± SEM for three independent enzyme purifications assayed in triplicate. B) Two-hybrid interaction of GBD-GKRP with GAD-GK or GAD-GK(p.Ala449Thr) mutant. Yeast strain Y187 was used, and fusion proteins were expressed from pGBKT7 and pACTII derivatives. Values are means ± SEM from ß-galactosidase activity of six independent transformants. In control experiments, GBD-GKRP did not interact with GAD and GAD-GK did not interact with GBD [22] .

    Journal: PLoS ONE

    Article Title: Functional Characterization of MODY2 Mutations Highlights the Importance of the Fine-Tuning of Glucokinase and Its Role in Glucose Sensing

    doi: 10.1371/journal.pone.0030518

    Figure Lengend Snippet: Effect of mutation p.Ala449Thr on the GK interaction with GKRP. A) Inhibition of glucokinase activity by human GKRP. Enzyme activity was measured at 5 mM glucose as described in Material and Methods , in the presence of 10 µM S6P (left panel) or 0.2 mM F1P (right panel). Results are means ± SEM for three independent enzyme purifications assayed in triplicate. B) Two-hybrid interaction of GBD-GKRP with GAD-GK or GAD-GK(p.Ala449Thr) mutant. Yeast strain Y187 was used, and fusion proteins were expressed from pGBKT7 and pACTII derivatives. Values are means ± SEM from ß-galactosidase activity of six independent transformants. In control experiments, GBD-GKRP did not interact with GAD and GAD-GK did not interact with GBD [22] .

    Article Snippet: Plasmids encoding a GKRP fusion protein to the Gal4 Binding Domain (GBD) in the pGBKT7 vector (Clontech) and a fusion of GK to the Gal4 Activating Domain (GAD) in the pACTII (Clontech), have been described previously .

    Techniques: Mutagenesis, Inhibition, Activity Assay

    Effect of mutation p.Ala449Thr on the GK interaction with GKRP. A) Inhibition of glucokinase activity by human GKRP. Enzyme activity was measured at 5 mM glucose as described in Material and Methods , in the presence of 10 µM S6P (left panel) or 0.2 mM F1P (right panel). Results are means ± SEM for three independent enzyme purifications assayed in triplicate. B) Two-hybrid interaction of GBD-GKRP with GAD-GK or GAD-GK(p.Ala449Thr) mutant. Yeast strain Y187 was used, and fusion proteins were expressed from pGBKT7 and pACTII derivatives. Values are means ± SEM from ß-galactosidase activity of six independent transformants. In control experiments, GBD-GKRP did not interact with GAD and GAD-GK did not interact with GBD [22] .

    Journal: PLoS ONE

    Article Title: Functional Characterization of MODY2 Mutations Highlights the Importance of the Fine-Tuning of Glucokinase and Its Role in Glucose Sensing

    doi: 10.1371/journal.pone.0030518

    Figure Lengend Snippet: Effect of mutation p.Ala449Thr on the GK interaction with GKRP. A) Inhibition of glucokinase activity by human GKRP. Enzyme activity was measured at 5 mM glucose as described in Material and Methods , in the presence of 10 µM S6P (left panel) or 0.2 mM F1P (right panel). Results are means ± SEM for three independent enzyme purifications assayed in triplicate. B) Two-hybrid interaction of GBD-GKRP with GAD-GK or GAD-GK(p.Ala449Thr) mutant. Yeast strain Y187 was used, and fusion proteins were expressed from pGBKT7 and pACTII derivatives. Values are means ± SEM from ß-galactosidase activity of six independent transformants. In control experiments, GBD-GKRP did not interact with GAD and GAD-GK did not interact with GBD [22] .

    Article Snippet: Plasmids encoding a GKRP fusion protein to the Gal4 Binding Domain (GBD) in the pGBKT7 vector (Clontech) and a fusion of GK to the Gal4 Activating Domain (GAD) in the pACTII (Clontech), have been described previously .

    Techniques: Mutagenesis, Inhibition, Activity Assay

    E2E1, E2G1, E2J1, E2J2, and E2L3 interact with MuRF1 in the presence of telethonin (A) Telethonin does not interact with E2s. Y2H experiments were performed using telethonin as a bait to confirm that this protein cannot directly interact with the E2 enzymes used in this work. The empty vector and the vector containing the LT construct were used as negative controls against telethonin to estimate potential background level. Signals above ‘empty’ and ‘LT’ lanes were considered as positive. Colonies were plated on selective medium [ ‐LTH + Aureo + 3‐AT ] (Experimental section) and monitored during 21 days. LT, Large‐T antigen; Tele, telethonin. (B) Telethonin expression level in yeast varies a ccording to methionine (Met) concentration in the medium. BD‐Tele, fusion protein between the binding domain of GAL4 and telethonin; Tele, telethonin. (C) Densitometry analysis from the immunoblot presented in (B). (D) Yeast three‐hybrid (Y3H) experiments revealed E2E1, E2G1, E2J1, E2J2, and E2L3 as MuRF1 partners in the presence of telethonin. E2‐expressing yeasts were mated against strains expressing either MuRF1 alone or MuRF1 and telethonin. Colonies were plated on selective medium [ ‐LTH + Aureo + 3‐AT ] containing 134 mM Met. Results were observed at day 6. Three to four independent transformation experiments were performed and 11 to 32 colonies were analyzed for each E2.

    Journal: Journal of Cachexia, Sarcopenia and Muscle

    Article Title: A muscle‐specific MuRF1‐E2 network requires stabilization of MuRF1‐E2 complexes by telethonin, a newly identified substrate) A muscle‐specific MuRF1‐E2 network requires stabilization of MuRF1‐E2 complexes by telethonin, a newly identified substrate

    doi: 10.1002/jcsm.12249

    Figure Lengend Snippet: E2E1, E2G1, E2J1, E2J2, and E2L3 interact with MuRF1 in the presence of telethonin (A) Telethonin does not interact with E2s. Y2H experiments were performed using telethonin as a bait to confirm that this protein cannot directly interact with the E2 enzymes used in this work. The empty vector and the vector containing the LT construct were used as negative controls against telethonin to estimate potential background level. Signals above ‘empty’ and ‘LT’ lanes were considered as positive. Colonies were plated on selective medium [ ‐LTH + Aureo + 3‐AT ] (Experimental section) and monitored during 21 days. LT, Large‐T antigen; Tele, telethonin. (B) Telethonin expression level in yeast varies a ccording to methionine (Met) concentration in the medium. BD‐Tele, fusion protein between the binding domain of GAL4 and telethonin; Tele, telethonin. (C) Densitometry analysis from the immunoblot presented in (B). (D) Yeast three‐hybrid (Y3H) experiments revealed E2E1, E2G1, E2J1, E2J2, and E2L3 as MuRF1 partners in the presence of telethonin. E2‐expressing yeasts were mated against strains expressing either MuRF1 alone or MuRF1 and telethonin. Colonies were plated on selective medium [ ‐LTH + Aureo + 3‐AT ] containing 134 mM Met. Results were observed at day 6. Three to four independent transformation experiments were performed and 11 to 32 colonies were analyzed for each E2.

    Article Snippet: Please refer to Table for other E2 enzymes nomenclature. cDNAs were cloned in yeast two‐hybrid vectors pGADT7, producing a fusion protein with the activation domain (AD) of GAL4 transcription factor, or in pGBKT7 and pBridge vectors, producing a fusion protein with the binding domain (BD) of GAL4 (Clontech).

    Techniques: Plasmid Preparation, Construct, Expressing, Concentration Assay, Binding Assay, Transformation Assay

    N protein of SARS-CoV associates with EF1α. (A) Interaction of N protein with EF1α was revealed by a yeast two-hybrid system. Three transformants containing pGBKT7-N and pACT2-EF1α(291-463) (left panel) together with both positive (upper right) and negative (lower right) controls were inoculated on SD-Leu − Trp − His − Ade − -X-α-Gal plates. Transformants containing pGADT7-T and pGBKT7-p53 were used as a positive control, and transformants containing pGADT7-T and a pGBKT7-Lam were employed as a negative control. (B) Lysates from 293T cells transfected with Flag-EF1α or Flag and GFP-N or GFP plasmids were subjected to immunoprecipitation with anti-Flag antibody and analyzed by immunoblotting with anti-GFP or anti-Flag antibody. (C) Lysates from 293T cells transfected with Flag-N or Flag plasmid were subjected to immunoprecipitation with anti-Flag and analyzed by immunoblotting with anti-EF1α antibody. (D) Lysates from 293T cells transfected with GFP-N or the indicated GFP-N truncation forms and Flag-EF1α were subjected to immunoprecipitation with anti-Flag and subsequently analyzed by immunoblotting with anti-GFP or anti-Flag antibody. IP, immunoprecipitation; IB, immunoblotting; P/N, positive/negative.

    Journal: Journal of Virology

    Article Title: The Nucleocapsid Protein of Severe Acute Respiratory Syndrome Coronavirus Inhibits Cell Cytokinesis and Proliferation by Interacting with Translation Elongation Factor 1α

    doi: 10.1128/JVI.00133-08

    Figure Lengend Snippet: N protein of SARS-CoV associates with EF1α. (A) Interaction of N protein with EF1α was revealed by a yeast two-hybrid system. Three transformants containing pGBKT7-N and pACT2-EF1α(291-463) (left panel) together with both positive (upper right) and negative (lower right) controls were inoculated on SD-Leu − Trp − His − Ade − -X-α-Gal plates. Transformants containing pGADT7-T and pGBKT7-p53 were used as a positive control, and transformants containing pGADT7-T and a pGBKT7-Lam were employed as a negative control. (B) Lysates from 293T cells transfected with Flag-EF1α or Flag and GFP-N or GFP plasmids were subjected to immunoprecipitation with anti-Flag antibody and analyzed by immunoblotting with anti-GFP or anti-Flag antibody. (C) Lysates from 293T cells transfected with Flag-N or Flag plasmid were subjected to immunoprecipitation with anti-Flag and analyzed by immunoblotting with anti-EF1α antibody. (D) Lysates from 293T cells transfected with GFP-N or the indicated GFP-N truncation forms and Flag-EF1α were subjected to immunoprecipitation with anti-Flag and subsequently analyzed by immunoblotting with anti-GFP or anti-Flag antibody. IP, immunoprecipitation; IB, immunoblotting; P/N, positive/negative.

    Article Snippet: The strain containing pGBKT7-N was further transformed with a fetal liver cDNA library cloned in fusion with the GAL4 activation domain using the pACT2 vector (Clontech).

    Techniques: Positive Control, Laser Capture Microdissection, Negative Control, Transfection, Immunoprecipitation, Plasmid Preparation

    FANCC directly interacts with UNC5A in yeasts. Yeast two-hybrid assays were performed with various UNC5A constructs that included the Y2H clone coding for the C-terminal death domain (DEATH) and a part of intron 14 (shown in blue), UNC5A intracellular domain (UNC5A ICD ), the UNC5A C-terminus (UNC5A DD ) and the death-domain deletion mutant (UNC5A ΔDD ). The yeast strain AH109 was co-transformed with UNC5A constructs along with FANCC constructs as indicated. The plus sign (+) indicate a positive interaction. Negative controls were performed using empty vectors (Empty), and positive controls were performed using pGBKT7/p53 and pGADT7/SV40 T-antigen from Clontech (data not shown). Yeast two-hybrid assays were performed at least three times (independent transformation) with 3 to 4 clones each.

    Journal: PLoS ONE

    Article Title: The Fanconi Anemia Group C Protein Interacts with Uncoordinated 5A and Delays Apoptosis

    doi: 10.1371/journal.pone.0092811

    Figure Lengend Snippet: FANCC directly interacts with UNC5A in yeasts. Yeast two-hybrid assays were performed with various UNC5A constructs that included the Y2H clone coding for the C-terminal death domain (DEATH) and a part of intron 14 (shown in blue), UNC5A intracellular domain (UNC5A ICD ), the UNC5A C-terminus (UNC5A DD ) and the death-domain deletion mutant (UNC5A ΔDD ). The yeast strain AH109 was co-transformed with UNC5A constructs along with FANCC constructs as indicated. The plus sign (+) indicate a positive interaction. Negative controls were performed using empty vectors (Empty), and positive controls were performed using pGBKT7/p53 and pGADT7/SV40 T-antigen from Clontech (data not shown). Yeast two-hybrid assays were performed at least three times (independent transformation) with 3 to 4 clones each.

    Article Snippet: Plasmids and DNA constructs The N-terminus of FANCC, which spans from nucleotides 256 to 1175 and encompasses amino acids from the start codon to the cleavage site , was cloned into the pGBKT7 and pGADT7 yeast vectors (Clontech Laboratories Inc., Mountain View, CA) by fusion to the Gal4-DNA binding or DNA-activating domain, and into the pEGFP plasmids (pGBKFANCC1-306 , pGADFANCC1-306 , pEGFPFANCC1-306 ).

    Techniques: Construct, Mutagenesis, Transformation Assay, Clone Assay