Engineering Oncohistone Mutants for Study

Double-stranded cDNAs encoding carboxyl-terminal FLAG epitope-tagged human wild-type and site-mutant histones H2B, H2A, H3, and H4 were synthesized as gene blocks (Integrated DNA Technologies), and then cloned individually into pCDH-EF1α-MCS-IRES-Puro lentiviral expression vector using Gibson assembly (NEB, E2621). Twenty-five individual glutamate (Glu, E) and aspartate (Asp, D) oncohistone sites (3 (link)) were mutated into alanine (Ala, A). H2B-D51 and H4-D68 were also mutated into asparagine (Asn, N).

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Huang D., Camacho C.V., Martire S., Nagari A., Setlem R., Gong X., Edwards A.D., Chiu S.P., Banaszynski L.A, & Kraus W.L. (2022). Oncohistone Mutations Occur at Functional Sites of Regulatory ADP-ribosylation. Cancer research, 82(13), 2361-2377.

Publication 2022

gene blocks

Alanine Aspartate Cdnas Epitope Gene Glutamate Human Ires Vector

Corresponding Organization : Union Hospital

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Variable analysis

independent variables

Twenty-five individual glutamate (Glu, E) and aspartate (Asp, D) oncohistone sites (3 (link)) were mutated into alanine (Ala, A).
H2B-D51 and H4-D68 were also mutated into asparagine (Asn, N).

dependent variables

Not explicitly mentioned.

control variables

Double-stranded cDNAs encoding carboxyl-terminal FLAG epitope-tagged human wild-type histones H2B, H2A, H3, and H4 were synthesized as gene blocks (Integrated DNA Technologies), and then cloned individually into pCDH-EF1α-MCS-IRES-Puro lentiviral expression vector using Gibson assembly (NEB, E2621).

controls

Wild-type histones H2B, H2A, H3, and H4 were used as positive controls.
No negative controls were explicitly mentioned.

Annotations

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