Northern Blotting Analysis of gDNA

Northern blotting was performed as described previously (37 (link)). gDNA was used as PCR templates for northern blotting probes. Probes labeled with ³²P-dCTP were generated from PCR templates using the Decaprime II kit according to manufacturer's instructions (Invitrogen). RNA samples were run on an agarose-formaldehyde gel and then transferred to a nylon membrane. After hybridization, Northern blots were quantified by densitometry using ImageQuant TL v7.0 (GE). Transcript levels were determined relative to the SCR1 Control. Primers used for making Northern probes are listed in Table S6.

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Hegazy Y.A., Cloutier S.C., Utturkar S.M., Das S, & Tran E.J. (2023). The genomic region of the 3′ untranslated region (3′UTR) of PHO84, rather than the antisense RNA, promotes gene repression. Nucleic Acids Research, 51(15), 7900-7913.

Publication 2023

Decaprime II kit ImageQuant TL v7.0

Agarose Dctp Densitometry Formaldehyde Hybridization Membrane Northern blots Nylon Primers

Corresponding Organization :

Other organizations : Purdue University West Lafayette

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Variable analysis

independent variables

None explicitly mentioned

dependent variables

Transcript levels

control variables

GDNA used as PCR templates for northern blotting probes
RNA samples run on an agarose-formaldehyde gel
RNA samples transferred to a nylon membrane
SCR1 used as control for determining transcript levels

positive controls

None explicitly mentioned

negative controls

None explicitly mentioned

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