WCRB2014 POSTER PRESENTATIONS (1) (335 abstracts)
Analysis of marsupial ZP: molecular and phylogenetic approach
C Moros Nicolás 1 , M J Izquierdo Rico 1 , W V Holt 2 , P Chevret 3 , M López-Bejar 4 , E Martínez-Nevado 5 , J Ballesta 1 & M Avilés 1
1University of Murcia, Murcia, Spain; 2Institute of Zoology, London, UK; 3Université Claude Bernard Lyon 1, Lyon, France; 4Universitat Autònoma de Barcelona, Barcelona, Spain; 5Zoo-Aquarium Madrid, Madrid, Spain.
Introduction: The zona pellucida (ZP) is an extracellular matrix that surrounds mammalian oocytes. Marsupial ZP has been considered formed by three proteins. However, the recent description of four glycoproteins (ZP1, ZP2, ZP3, and ZP4) in the ZP of some species of eutherian (rat, human, hamster, rabbit) suggests the need for a reanalysis of the ZP composition in marsupials.
Material and methods: Complementary DNA was obtained from ovaries from one Australasian marsupial (Bennett’s wallaby) and genomic DNA was obtained from one Australasian marsupial (koala) and two South American marsupials (common opossum and Pinheiro’s slender opossum). Specific primers for ZP1, ZP2, ZP3, and ZP4 were designed for PCR analysis. Different amplicons were obtained and automatically sequenced.
Results and discussion: In wallaby, PCR amplifications resulted in the complete amplification of the open reading frame (ORF) of ZP1 and ZP4 and partial amplifications of ZP2 and ZP3. In opossums the analysis of the amplicons obtained indicated that ZP4 present stop codons along its sequence. In Koala, ZP4 is free of stop codons. Wallaby ZP expresses four genes. Koala ZP expresses a functional ZP4, while in common opossum and Pinheiro’s slender opossum ZP4 is not functional. These results strongly suggest the presence of a ZP4 pseudogene in the Didelphimorphia order, demonstrating that ZP composition is different in Australasian and South American marsupials.
This work is supported by MINECO (AGL2012-40180-C03-02), European Commission (FEDER/ERDF) and Fundación Séneca (0452/GERM/06).
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