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      Expression of luteinizing hormone receptor during development of bovine fetal ovary


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          Steroids and gonadotrophins are essential for the regulation of late stages of preantral development and antral follicular development. Although the luteinizing hormone receptor (LHCGR) has been detected in the preantral follicles of rats, rabbits, and pigs, its expression, in bovine fetal ovary, has not been demonstrated. Based on this, we aimed to investigate the expression of the LHCGR and LHCGR mRNA binding protein ( LRBP), as well as, to quantify bta-miR-222 (a regulatory microRNA of the LHCGR gene) during the development of bovine fetal ovary. In summary, LHCGR expression was observed in the preantral follicle in bovine fetal ovary, from oogonias to primordial, primary and secondary stages, and the mRNA abundance was lower on day 150 than day 60. However, the mRNA abundance of LRBP followed the opposite pattern. Similar to LRBP, the abundance of bta-miR-222 was higher on day 150 than day 60 or 90 of gestation. The LHCGR protein was detected in oogonia, primordial, primary, and secondary follicles. Moreover, both oocytes and granulosa cells showed positive immunostaining for LHCGR. In conclusion, we suggest the involvement of LHCGR/LRBP/bta-mir222 with mechanisms related to the development of preantral follicles in cattle.

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          A new mathematical model for relative quantification in real-time RT-PCR.

          M. Pfaffl (2001)
          Use of the real-time polymerase chain reaction (PCR) to amplify cDNA products reverse transcribed from mRNA is on the way to becoming a routine tool in molecular biology to study low abundance gene expression. Real-time PCR is easy to perform, provides the necessary accuracy and produces reliable as well as rapid quantification results. But accurate quantification of nucleic acids requires a reproducible methodology and an adequate mathematical model for data analysis. This study enters into the particular topics of the relative quantification in real-time RT-PCR of a target gene transcript in comparison to a reference gene transcript. Therefore, a new mathematical model is presented. The relative expression ratio is calculated only from the real-time PCR efficiencies and the crossing point deviation of an unknown sample versus a control. This model needs no calibration curve. Control levels were included in the model to standardise each reaction run with respect to RNA integrity, sample loading and inter-PCR variations. High accuracy and reproducibility (<2.5% variation) were reached in LightCycler PCR using the established mathematical model.
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            Assumption-free analysis of quantitative real-time polymerase chain reaction (PCR) data.

            Quantification of mRNAs using real-time polymerase chain reaction (PCR) by monitoring the product formation with the fluorescent dye SYBR Green I is being extensively used in neurosciences, developmental biology, and medical diagnostics. Most PCR data analysis procedures assume that the PCR efficiency for the amplicon of interest is constant or even, in the case of the comparative C(t) method, equal to 2. The latter method already leads to a 4-fold error when the PCR efficiencies vary over just a 0.04 range. PCR efficiencies of amplicons are usually calculated from standard curves based on either known RNA inputs or on dilution series of a reference cDNA sample. In this paper we show that the first approach can lead to PCR efficiencies that vary over a 0.2 range, whereas the second approach may be off by 0.26. Therefore, we propose linear regression on the Log(fluorescence) per cycle number data as an assumption-free method to calculate starting concentrations of mRNAs and PCR efficiencies for each sample. A computer program to perform this calculation is available on request (e-mail: bioinfo@amc.uva.nl; subject: LinRegPCR).
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              Oocyte control of ovarian follicular development and function in mammals.

              A new perspective on ovarian follicular development has emerged over the last decade. Whereas the oocyte was previously considered only a passive recipient of developmental signals from oocyte-associated granulosa cells, it is now clear that communication between oocytes and granulosa cells is bidirectional. A complex interplay of regulatory factors governs the development of both types of cell. This interplay is essential not only for oocyte development but also for follicular development, beginning with the initial assembly of the primordial follicle and continuing throughout ovulation. The existence of an oocyte-granulosa cell regulatory loop, essential for normal follicular differentiation as well as for the production of an oocyte competent to undergo fertilization and embryogenesis, is proposed. Although gonadotrophins are essential for driving the differentiation of granulosa cell phenotypes, within its sphere of influence, the oocyte is probably the dominant factor determining the direction of differentiation and the function of the granulosa cells associated with it.

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                Role: conceptualizationRole: funding acquisitionRole: investigationRole: writingRole: original draftRole: project administrationRole: data curation
                Role: writingRole: review & editingRole: formal analysis
                Role: data curationRole: writingRole: review & editing
                Role: investigationRole: data curationRole: writingRole: review & editing
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                Role: conceptualizationRole: investigationRole: project administrationRole: writingRole: original draftRole: writingRole: review & editing
                Anim Reprod
                Anim Reprod
                Animal Reproduction
                Colégio Brasileiro de Reprodução Animal
                15 April 2024
                : 21
                : 1
                : e20230112
                [1 ] originalUniversidade do Oeste Paulista, Presidente Prudente, SP, Brasil
                [2 ] originalDepartamento de Farmacologia, Instituto de Biociências, Universidade Estadual Paulista, Botucatu, SP, Brasil
                Author notes
                [* ]Corresponding author: castilho.anthony@ 123456gmail.com

                Conflicts of interest: The authors have no conflict of interest to declare.

                Author information
                arAO20230112_EN 00211

                Copyright © The Author(s). This is an Open Access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

                : 15 June 2023
                : 19 February 2024
                Page count
                Figures: 3, Tables: 0, Equations: 0, References: 36
                Funded by: FAPESP
                Award ID: #2013/11480-3,
                Award ID: #2015/04505-5
                Original Article

                lhcgr,preantral follicle,steroidogenesis,micrornas
                lhcgr, preantral follicle, steroidogenesis, micrornas


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