Posts Tagged ‘Estrogen

Northern Blotting and ISH Analyses Porcine uterine ER mRNA was detected by Northern blot and ISH analyses using [a-32P]UTP- and [35S]UTP-labeled antisense cRNA probes generated from 0ER8 cDNA template. Using the [a-32P]UTP-labeled antisense ER cRNA probe, a single major transcript of approximately 6.2 kilobases (kb) was detected by Northern blot analysis of total uterine RNA […]

Age-related changes in ER immunostaining intensity observed for GE, LE, and S are summarized in Figure 5. Nuclear staining was absent in endometrium from PND 0. However, by PND 15, with the appearance of endometrial glands, strong positive nuclear staining was observed in GE. In striking contrast, nuclear ER staining was effectively absent in LE […]

Western Blotting Results of Western blot analyses of uterine tissue protein from PND 0, 15, and 120 are shown in Figure 3. In tissue homogenate from PND 0, no specific signal of the expected size for ER protein (approximately 66 kDa) was detected. However, a specific positive ER signal of the expected size was observed as […]

Uterine Growth Effects of age and OVX at birth on uterine wet weight and endometrial thickness are summarized in Figure 1. Both uterine weight and endometrial thickness were affected by age and OVX (p < 0.01), and an interaction (p < 0.01) between age and OVX was detected for both of these dependent variables. Overall, […]

Slides of porcine endometrium developed to display in situ-hybridized ER antisense and sense cRNAs were examined at XlO, X20, and X40 by darkfield microscopy on the same microscope equipped with an Olympus darkfield condenser. Darkfield images were captured from a silicon intensified target (SIT) camera (model 68; Dage/MTI). Both brightfield and darkfield images were background […]

In Situ Hybridization In situ hybridization (ISH) was performed, using [35S]UTP-labeled (specific activity: 800 Ci/mmol; Amersham, Arlington Heights, IL) antisense and sense cRNA probes produced by in vitro transcription from the EcoRI-and BamHI-linearized 0ER8 cDNA template, with a MaxiScript kit (Ambion, Austin, TX). T7 and SP6 RNA polymerases were used to transcribe antisense and sense […]

Northern Blotting Northern hybridization was performed to confirm specificity of the ovine 0ER8 probe for porcine RNA. The sequence of this partial cDNA for the ovine ER shares significant homology with ERa of other species, but not with the recently cloned ER. Total cellular RNA was extracted from uteri using TRIzol Reagent (Life Technologies, Gaithersburg, […]

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