Source: An E. coli strain that carries a plasmid Mass Spectrometry: The mass of purified Exonuclease Assay: Incubation of a 50 µl Histone H3.2 encoding the cloned human histone H3.2 gene, Histone H3.2 Human, Recombinant is reaction containing 10 µg of Histone H3.2 Human, HIST2H3A or HIST2H3C. (Genbank accession 15257.09 Da as determined by ESI-TOF MS Recombinant with 1 µg of a mixture of single and Human, Recombinant number: BC130637) (Electrospray Ionization-Time of Flight Mass double-stranded [3H] E. coli DNA (200,000 cpm/ Spectrometry). The average mass calculated from µg) for 4 hours at 37°C released < 0.1% of the 1-800-632-7799 Supplied in: 20 mM Sodium Phosphate (pH 7.0), primary sequence is 15256.82 Da. This confirms total radioactivity. [email protected] 300 mM NaCl, 1 mM EDTA and 1 mM DTT. the protein identity of the histone. For a typical www.neb.com example of mass spectrometry data, please see Endonuclease Assay: Incubation of a 50 µl M2506S 002150417041 Note: The protein concentration (1 mg/ml, 66 µM) the product page at www.neb.com. reaction containing 10 µg of Histone H3.2 is calculated using the molar extinction coefficient Human, Recombinant with 1 µg of φX174 RF I for Histone H3.2 (3960) and its absorbance at N-terminal Protein Sequencing: Protein identity (supercoiled) plasmid DNA for 4 hours at 37°C M2506S was confirmed using Edman Degradation to resulted in < 5.0% conversion to RF II form 280 nm (4,5). 1.0 A280 units = 3.9 mg/ml 100 µg 1.0 mg/ml Lot: 0021504 sequence the intact protein. (nicked circle) as determined by agarose gel Synonym: Histone H3/m, H3/o electrophoresis. RECOMBINANT Store at –20°C Exp: 4/17 Enzyme Modification: SET7 Methyltransferase: Gene Synonym: H3F2, H3FM Description: Histone H3 combines with Histone After incubation of a 25 µl reaction for 10 minutes Protein Sequence: ARTKQTARKSTGGKAPRKQLA H4 to form the H3/H4 tetramer. Two H2A/H2B at 37°C, 1 unit of SET7 methyltransferase TKAARKSAPATGGVKKPHRYRPGTVALREIRRYQKS Quality Control Assays: heterodimers interact with an H3/H4 tetramer to transfers 20 pmols of methyl group to Histone TELLIRKLPFQRLVREIAQDFKTDLRFQSSAVMALQEA form the histone octamer (1,2). It is also modified SDS-PAGE: 0.5, 1.0, 2.0, 5.0, 10.0 µg of Histone H3.2 Human, Recombinant. SEAYLVGLFEDTNLCAIHAKRVTIMPKDIQLARRIRGE by various enzymes and can act as a substrate for H3.2 Human, Recombinant were loaded on a 10– RA (Genbank accession number: Q71DI3) Protease Assay: After incubation of 10 µg them. These modifications have been shown to be 20% Tris-Glycine SDS-PAGE gel and stained with of Histone H3.2 Human, Recombinant with a important in gene regulation. Coomassie Blue. The calculated molecular weight is 15256.82 Da. Its apparent molecular weight on standard mixture of proteins for 4 hours at 37°C, (see other side) Histone H3.2, an H3 variant that is found in all 10–20% Tris-Glycine SDS-PAGE gel is ~17 kDa. no proteolytic activity could be detected by SDS- eukaryotes except budding yeast, is replication de- For a typical example of gel image, please see the PAGE. pendent and is associated with gene silencing (3). product page at www.neb.com.

CERTIFICATE OF ANALYSIS

Source: An E. coli strain that carries a plasmid Mass Spectrometry: The mass of purified Exonuclease Assay: Incubation of a 50 µl Histone H3.2 encoding the cloned human histone H3.2 gene, Histone H3.2 Human, Recombinant is reaction containing 10 µg of Histone H3.2 Human, HIST2H3A or HIST2H3C. (Genbank accession 15257.09 Da as determined by ESI-TOF MS Recombinant with 1 µg of a mixture of single and Human, Recombinant number: BC130637) (Electrospray Ionization-Time of Flight Mass double-stranded [3H] E. coli DNA (200,000 cpm/ Spectrometry). The average mass calculated from µg) for 4 hours at 37°C released < 0.1% of the 1-800-632-7799 Supplied in: 20 mM Sodium Phosphate (pH 7.0), primary sequence is 15256.82 Da. This confirms total radioactivity. [email protected] 300 mM NaCl, 1 mM EDTA and 1 mM DTT. the protein identity of the histone. For a typical www.neb.com example of mass spectrometry data, please see Endonuclease Assay: Incubation of a 50 µl M2506S 002150417041 Note: The protein concentration (1 mg/ml, 66 µM) the product page at www.neb.com. reaction containing 10 µg of Histone H3.2 is calculated using the molar extinction coefficient Human, Recombinant with 1 µg of φX174 RF I for Histone H3.2 (3960) and its absorbance at N-terminal Protein Sequencing: Protein identity (supercoiled) plasmid DNA for 4 hours at 37°C M2506S was confirmed using Edman Degradation to resulted in < 5.0% conversion to RF II form 280 nm (4,5). 1.0 A280 units = 3.9 mg/ml 100 µg 1.0 mg/ml Lot: 0021504 sequence the intact protein. (nicked circle) as determined by agarose gel Synonym: Histone H3/m, H3/o electrophoresis. RECOMBINANT Store at –20°C Exp: 4/17 Enzyme Modification: SET7 Methyltransferase: Gene Synonym: H3F2, H3FM Description: Histone H3 combines with Histone After incubation of a 25 µl reaction for 10 minutes Protein Sequence: ARTKQTARKSTGGKAPRKQLA H4 to form the H3/H4 tetramer. Two H2A/H2B at 37°C, 1 unit of SET7 methyltransferase TKAARKSAPATGGVKKPHRYRPGTVALREIRRYQKS Quality Control Assays: heterodimers interact with an H3/H4 tetramer to transfers 20 pmols of methyl group to Histone TELLIRKLPFQRLVREIAQDFKTDLRFQSSAVMALQEA form the histone octamer (1,2). It is also modified SDS-PAGE: 0.5, 1.0, 2.0, 5.0, 10.0 µg of Histone H3.2 Human, Recombinant. SEAYLVGLFEDTNLCAIHAKRVTIMPKDIQLARRIRGE by various enzymes and can act as a substrate for H3.2 Human, Recombinant were loaded on a 10– RA (Genbank accession number: Q71DI3) Protease Assay: After incubation of 10 µg them. These modifications have been shown to be 20% Tris-Glycine SDS-PAGE gel and stained with of Histone H3.2 Human, Recombinant with a important in gene regulation. Coomassie Blue. The calculated molecular weight is 15256.82 Da. Its apparent molecular weight on standard mixture of proteins for 4 hours at 37°C, (see other side) Histone H3.2, an H3 variant that is found in all 10–20% Tris-Glycine SDS-PAGE gel is ~17 kDa. no proteolytic activity could be detected by SDS- eukaryotes except budding yeast, is replication de- For a typical example of gel image, please see the PAGE. pendent and is associated with gene silencing (3). product page at www.neb.com.

CERTIFICATE OF ANALYSIS References: 1. Kornberg, R.D. (1977) Annu. Rev. Biochem. 46, ISO 9001 ISO 14001 ISO 13485 931–954. Registered Registered Registered Quality Environmental Medical Devices 2. van Holde, K.E. (1989) Chromatin, 1–497. Management Management 3. Hake, S.B. et al (2006) J.Biol. Chem. 281, 559- New England Biolabs® is a registered trademark of New England Biolabs, Inc. 568. 4. Gill, S.C. and von Hippel, P.H. (1989) Anal. Bio- chem. 182, 319–326. 5. Pace, C.N. et al. (1995) Protein Science, 4, 2411–2423.

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References: 1. Kornberg, R.D. (1977) Annu. Rev. Biochem. 46, ISO 9001 ISO 14001 ISO 13485 931–954. Registered Registered Registered Quality Environmental Medical Devices 2. van Holde, K.E. (1989) Chromatin, 1–497. Management Management 3. Hake, S.B. et al (2006) J.Biol. Chem. 281, 559- New England Biolabs® is a registered trademark of New England Biolabs, Inc. 568. 4. Gill, S.C. and von Hippel, P.H. (1989) Anal. Bio- chem. 182, 319–326. 5. Pace, C.N. et al. (1995) Protein Science, 4, 2411–2423.

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