Chromogenic ISH analysis of miR-409-3p

The embryo (25 d) and extra-embryonic membranes (25 and 45 d) were retrieved by uterine lavage [29 (link)]. The tissue samples were fixed in 10% formaldehyde for 24 h, transferred to 70% methanol and paraffin embedded [29 (link)]. The expression of eca-miR-409-3p was investigated by chromogenic ISH using a dual digoxigenin (DIG)-labeled LNA™ probe specific to hsa-miR-409-3p (610701-360; miRCURY LNA™, Exiqon, Vedbaek, Denmark) as previously described [29 (link)] on 25 d embryos (n = 3) along with the extra-embryonic membranes. A dual DIG-labeled LNA™ probe specific to U6 snRNA (#699002-360; Exiqon) and a scrambled miRNA probe (#699003-360; Exiqon) were used as positive and negative controls, respectively.

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Dini P., El-Sheikh Ali H., Carossino M., C. Loux S., Esteller-Vico A., E. Scoggin K., Daels P, & A. Ball B. (2019). Expression Profile of the Chromosome 14 MicroRNA Cluster (C14MC) Ortholog in Equine Maternal Circulation throughout Pregnancy and Its Potential Implications. International Journal of Molecular Sciences, 20(24), 6285.

Publication 2019

miRCURY LNA™, scrambled miRNA probe

Chromogenic Digoxigenin Embryos Extra embryonic membranes Formaldehyde Hsa mir 409 Methanol Mirna Paraffin Tissue U6 snrna Uterine

Corresponding Organization : University of Kentucky

Other organizations : Ghent University, Mansoura University, Louisiana State University, University of Tennessee at Knoxville

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Variable analysis

independent variables

Gestational age of embryos and extra-embryonic membranes (25 days and 45 days)

dependent variables

Expression of eca-miR-409-3p

control variables

Tissue samples were fixed in 10% formaldehyde for 24 h, transferred to 70% methanol and paraffin embedded

controls

Positive control: Dual DIG-labeled LNA™ probe specific to U6 snRNA
Negative control: Scrambled miRNA probe

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