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| Author(s): |
Ana Paula Dini Andreote
Total Authors: 1
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| Document type: | Master's Dissertation |
| Press: | Piracicaba. |
| Institution: | Universidade de São Paulo (USP). Centro de Energia Nuclear na Agricultura (CENA/STB) |
| Defense date: | 2009-06-10 |
| Examining board members: |
Luiz Lehmann Coutinho;
Lucia Elvira Alvares;
Luis Felipe Prada e Silva
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| Advisor: | Luiz Lehmann Coutinho |
| Abstract | |
MicroRNAs (miRNAs) represent a class of small and noncoding RNAs, about 20-nucleotides long that negatively regulate gene expression posttranscriptionally. The regulatory action of these molecules is essential for the proper functioning of various biological processes, including the development and maintenance of skeletal muscles. To identify miRNAs that might be important for the skeletal muscle development, we constructed a miRNA library from pectoral skeletal muscle of 21º days after birth chickens. A total of 576 clones were sequenced and these sequences were collapsed into 98 clusters. Sequence analysis identified 47 small RNAs that show significant similarities with published miRNAs, 30 with others noncoding RNAs and six sequences clusters could be identified as potentially novel miRNAs. These data may support subsequent functional studies aimed at understanding the function performed by each of these molecules in skeletal muscle of chicken. To further associate the miRNA presence with the gene expression in the controlling of myogenesis the expression patterns of tree miRNAs identified in the library (miR-125b, miR-221 e miR-206) were analyzed. These miRNAs are involved in the balance between proliferation and differentiation mechanisms that control myogenesis. The expressions of these miRNAs were measured using quantitative RT-PCR. We analyzed samples from two embryos stages (9 and 17 days) and one adult stage (21º days after birth) in two chicken lines with different potential to growth and gain of muscle mass. There were no significant differences between the lines about these miRNAs expression. But, we could predict an overview of these miRNAs expressions during the muscle development of chicken, which allowed inferences about the physiologic and morphologic conditions of the muscles cells in each analyzed stage. Also, to further associate the miRNAs results to variations in the target genes expressions, we analyzed the expression of three genes: SRF, Fstl e Pola1. The first one is target of miR-133a (not analyzed due to methodological problems), and the others are target of miR-206. We analyzed by quantitative RT-PCR different stages and two chicken lines. It was possible to observe, only in the earlier stages, a relationship between the miR-206 expression and the Pola1 and Fstl1 expression, with a consistency between the increased expression of mir-206 and decrease in expression of Pola1 and Fstl1. The determination of the profile of these three gene expressions during the muscle development allowed inferences about the action of these molecules in the balance between proliferation and differentiation process in chicken strains for broiler and layer (AU) | |