Structured Review

Millipore 42a 44a
42a 44a, supplied by Millipore, used in various techniques. Bioz Stars score: 79/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/42a 44a/product/Millipore
Average 79 stars, based on 1 article reviews
Price from $9.99 to $1999.99
42a 44a - by Bioz Stars, 2020-01
79/100 stars

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Centrifugation:

Article Title: Phosphorylation of protein kinase C sites Ser42/44 decreases Ca2+-sensitivity and blunts enhanced length-dependent activation in response to protein kinase A in human cardiomyocytes
Article Snippet: Briefly, three different cTnI forms were made via site-directed mutations of Ser42/44 and Ser23/24 into aspartic acid (D) to mimic phosphorylation or into alanine (A) to mimic dephosphorylation: 42D/44D, 42A/44A and 23D/24D. cDNA encoding human cardiac isoforms (troponin C (cTnC), myc-tag labeled cTnT (cTnT-myc), cTnI wild-type, and cTnI mutants) were transformed in E. coli Rosetta2 and cultured under carbenicillin/chloramphenicol selection in Overnight Express TB medium (EMD Biosciences). .. Cultures were harvested by centrifugation, re-suspended in phosphate buffered saline (PBS), and centrifuged at 10000xg.

Selection:

Article Title: Phosphorylation of protein kinase C sites Ser42/44 decreases Ca2+-sensitivity and blunts enhanced length-dependent activation in response to protein kinase A in human cardiomyocytes
Article Snippet: .. Briefly, three different cTnI forms were made via site-directed mutations of Ser42/44 and Ser23/24 into aspartic acid (D) to mimic phosphorylation or into alanine (A) to mimic dephosphorylation: 42D/44D, 42A/44A and 23D/24D. cDNA encoding human cardiac isoforms (troponin C (cTnC), myc-tag labeled cTnT (cTnT-myc), cTnI wild-type, and cTnI mutants) were transformed in E. coli Rosetta2 and cultured under carbenicillin/chloramphenicol selection in Overnight Express TB medium (EMD Biosciences). .. Cultures were harvested by centrifugation, re-suspended in phosphate buffered saline (PBS), and centrifuged at 10000xg.

Expressing:

Article Title: Phosphorylation of protein kinase C sites Ser42/44 decreases Ca2+-sensitivity and blunts enhanced length-dependent activation in response to protein kinase A in human cardiomyocytes
Article Snippet: Paragraph title: Expression and purification of recombinant troponin subunits ... Briefly, three different cTnI forms were made via site-directed mutations of Ser42/44 and Ser23/24 into aspartic acid (D) to mimic phosphorylation or into alanine (A) to mimic dephosphorylation: 42D/44D, 42A/44A and 23D/24D. cDNA encoding human cardiac isoforms (troponin C (cTnC), myc-tag labeled cTnT (cTnT-myc), cTnI wild-type, and cTnI mutants) were transformed in E. coli Rosetta2 and cultured under carbenicillin/chloramphenicol selection in Overnight Express TB medium (EMD Biosciences).

Chromatography:

Article Title: Phosphorylation of protein kinase C sites Ser42/44 decreases Ca2+-sensitivity and blunts enhanced length-dependent activation in response to protein kinase A in human cardiomyocytes
Article Snippet: Briefly, three different cTnI forms were made via site-directed mutations of Ser42/44 and Ser23/24 into aspartic acid (D) to mimic phosphorylation or into alanine (A) to mimic dephosphorylation: 42D/44D, 42A/44A and 23D/24D. cDNA encoding human cardiac isoforms (troponin C (cTnC), myc-tag labeled cTnT (cTnT-myc), cTnI wild-type, and cTnI mutants) were transformed in E. coli Rosetta2 and cultured under carbenicillin/chloramphenicol selection in Overnight Express TB medium (EMD Biosciences). .. Fractions containing equal ratios of cTnT, cTnC and cTnI subunits were pooled, subsequently purified by AKTA-FPLC chromatography using a Resource Q to remove residual uncomplexed troponin subunit and finally dialyzed against 10 mM imidazole, 200 mM KCl, 5 mM MgCl2 , 2.5 mM EGTA, 1 mM DTT, 0.1 mM PMSF (pH 6.9; twice, 1L each) prior to concentrating the complexes to a final concentration > 2 mg/mL by centrifugation using Centriprep YM-10 centrifugal filters (Millipore).

Cell Culture:

Article Title: Phosphorylation of protein kinase C sites Ser42/44 decreases Ca2+-sensitivity and blunts enhanced length-dependent activation in response to protein kinase A in human cardiomyocytes
Article Snippet: .. Briefly, three different cTnI forms were made via site-directed mutations of Ser42/44 and Ser23/24 into aspartic acid (D) to mimic phosphorylation or into alanine (A) to mimic dephosphorylation: 42D/44D, 42A/44A and 23D/24D. cDNA encoding human cardiac isoforms (troponin C (cTnC), myc-tag labeled cTnT (cTnT-myc), cTnI wild-type, and cTnI mutants) were transformed in E. coli Rosetta2 and cultured under carbenicillin/chloramphenicol selection in Overnight Express TB medium (EMD Biosciences). .. Cultures were harvested by centrifugation, re-suspended in phosphate buffered saline (PBS), and centrifuged at 10000xg.

Purification:

Article Title: Phosphorylation of protein kinase C sites Ser42/44 decreases Ca2+-sensitivity and blunts enhanced length-dependent activation in response to protein kinase A in human cardiomyocytes
Article Snippet: Paragraph title: Expression and purification of recombinant troponin subunits ... Briefly, three different cTnI forms were made via site-directed mutations of Ser42/44 and Ser23/24 into aspartic acid (D) to mimic phosphorylation or into alanine (A) to mimic dephosphorylation: 42D/44D, 42A/44A and 23D/24D. cDNA encoding human cardiac isoforms (troponin C (cTnC), myc-tag labeled cTnT (cTnT-myc), cTnI wild-type, and cTnI mutants) were transformed in E. coli Rosetta2 and cultured under carbenicillin/chloramphenicol selection in Overnight Express TB medium (EMD Biosciences).

Produced:

Article Title: Phosphorylation of protein kinase C sites Ser42/44 decreases Ca2+-sensitivity and blunts enhanced length-dependent activation in response to protein kinase A in human cardiomyocytes
Article Snippet: Recombinant human troponin complex was produced as described in detail previously. .. Briefly, three different cTnI forms were made via site-directed mutations of Ser42/44 and Ser23/24 into aspartic acid (D) to mimic phosphorylation or into alanine (A) to mimic dephosphorylation: 42D/44D, 42A/44A and 23D/24D. cDNA encoding human cardiac isoforms (troponin C (cTnC), myc-tag labeled cTnT (cTnT-myc), cTnI wild-type, and cTnI mutants) were transformed in E. coli Rosetta2 and cultured under carbenicillin/chloramphenicol selection in Overnight Express TB medium (EMD Biosciences).

Concentration Assay:

Article Title: Phosphorylation of protein kinase C sites Ser42/44 decreases Ca2+-sensitivity and blunts enhanced length-dependent activation in response to protein kinase A in human cardiomyocytes
Article Snippet: Briefly, three different cTnI forms were made via site-directed mutations of Ser42/44 and Ser23/24 into aspartic acid (D) to mimic phosphorylation or into alanine (A) to mimic dephosphorylation: 42D/44D, 42A/44A and 23D/24D. cDNA encoding human cardiac isoforms (troponin C (cTnC), myc-tag labeled cTnT (cTnT-myc), cTnI wild-type, and cTnI mutants) were transformed in E. coli Rosetta2 and cultured under carbenicillin/chloramphenicol selection in Overnight Express TB medium (EMD Biosciences). .. Fractions containing equal ratios of cTnT, cTnC and cTnI subunits were pooled, subsequently purified by AKTA-FPLC chromatography using a Resource Q to remove residual uncomplexed troponin subunit and finally dialyzed against 10 mM imidazole, 200 mM KCl, 5 mM MgCl2 , 2.5 mM EGTA, 1 mM DTT, 0.1 mM PMSF (pH 6.9; twice, 1L each) prior to concentrating the complexes to a final concentration > 2 mg/mL by centrifugation using Centriprep YM-10 centrifugal filters (Millipore).

Labeling:

Article Title: Phosphorylation of protein kinase C sites Ser42/44 decreases Ca2+-sensitivity and blunts enhanced length-dependent activation in response to protein kinase A in human cardiomyocytes
Article Snippet: .. Briefly, three different cTnI forms were made via site-directed mutations of Ser42/44 and Ser23/24 into aspartic acid (D) to mimic phosphorylation or into alanine (A) to mimic dephosphorylation: 42D/44D, 42A/44A and 23D/24D. cDNA encoding human cardiac isoforms (troponin C (cTnC), myc-tag labeled cTnT (cTnT-myc), cTnI wild-type, and cTnI mutants) were transformed in E. coli Rosetta2 and cultured under carbenicillin/chloramphenicol selection in Overnight Express TB medium (EMD Biosciences). .. Cultures were harvested by centrifugation, re-suspended in phosphate buffered saline (PBS), and centrifuged at 10000xg.

De-Phosphorylation Assay:

Article Title: Phosphorylation of protein kinase C sites Ser42/44 decreases Ca2+-sensitivity and blunts enhanced length-dependent activation in response to protein kinase A in human cardiomyocytes
Article Snippet: .. Briefly, three different cTnI forms were made via site-directed mutations of Ser42/44 and Ser23/24 into aspartic acid (D) to mimic phosphorylation or into alanine (A) to mimic dephosphorylation: 42D/44D, 42A/44A and 23D/24D. cDNA encoding human cardiac isoforms (troponin C (cTnC), myc-tag labeled cTnT (cTnT-myc), cTnI wild-type, and cTnI mutants) were transformed in E. coli Rosetta2 and cultured under carbenicillin/chloramphenicol selection in Overnight Express TB medium (EMD Biosciences). .. Cultures were harvested by centrifugation, re-suspended in phosphate buffered saline (PBS), and centrifuged at 10000xg.

Transformation Assay:

Article Title: Phosphorylation of protein kinase C sites Ser42/44 decreases Ca2+-sensitivity and blunts enhanced length-dependent activation in response to protein kinase A in human cardiomyocytes
Article Snippet: .. Briefly, three different cTnI forms were made via site-directed mutations of Ser42/44 and Ser23/24 into aspartic acid (D) to mimic phosphorylation or into alanine (A) to mimic dephosphorylation: 42D/44D, 42A/44A and 23D/24D. cDNA encoding human cardiac isoforms (troponin C (cTnC), myc-tag labeled cTnT (cTnT-myc), cTnI wild-type, and cTnI mutants) were transformed in E. coli Rosetta2 and cultured under carbenicillin/chloramphenicol selection in Overnight Express TB medium (EMD Biosciences). .. Cultures were harvested by centrifugation, re-suspended in phosphate buffered saline (PBS), and centrifuged at 10000xg.

Recombinant:

Article Title: Phosphorylation of protein kinase C sites Ser42/44 decreases Ca2+-sensitivity and blunts enhanced length-dependent activation in response to protein kinase A in human cardiomyocytes
Article Snippet: Paragraph title: Expression and purification of recombinant troponin subunits ... Briefly, three different cTnI forms were made via site-directed mutations of Ser42/44 and Ser23/24 into aspartic acid (D) to mimic phosphorylation or into alanine (A) to mimic dephosphorylation: 42D/44D, 42A/44A and 23D/24D. cDNA encoding human cardiac isoforms (troponin C (cTnC), myc-tag labeled cTnT (cTnT-myc), cTnI wild-type, and cTnI mutants) were transformed in E. coli Rosetta2 and cultured under carbenicillin/chloramphenicol selection in Overnight Express TB medium (EMD Biosciences).

Fast Protein Liquid Chromatography:

Article Title: Phosphorylation of protein kinase C sites Ser42/44 decreases Ca2+-sensitivity and blunts enhanced length-dependent activation in response to protein kinase A in human cardiomyocytes
Article Snippet: Briefly, three different cTnI forms were made via site-directed mutations of Ser42/44 and Ser23/24 into aspartic acid (D) to mimic phosphorylation or into alanine (A) to mimic dephosphorylation: 42D/44D, 42A/44A and 23D/24D. cDNA encoding human cardiac isoforms (troponin C (cTnC), myc-tag labeled cTnT (cTnT-myc), cTnI wild-type, and cTnI mutants) were transformed in E. coli Rosetta2 and cultured under carbenicillin/chloramphenicol selection in Overnight Express TB medium (EMD Biosciences). .. Troponin subunits were purified using fast protein liquid chromatography (AKTA-FPLC System Amersham Biosciences) essentially as described previously.

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    Millipore 42a 44a
    42a 44a, supplied by Millipore, used in various techniques. Bioz Stars score: 79/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/42a 44a/product/Millipore
    Average 79 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    42a 44a - by Bioz Stars, 2020-01
    79/100 stars
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