Isolation and characterization of genes functionally involved in ovarian development of the giant tiger shrimp Penaeus monodon by suppression subtractive hybridization (SSH)

Rachanimuk Preechaphol,Piamsak Menasveta,Sirawut Klinbunga,Bavornlak Khamnamtongand

doi:10.1590/s1415-47572010000400014

Rachanimuk Preechaphol, Piamsak Menasveta + Show 2 more

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Suppression subtractive hybridization (SSH) libraries between cDNA in stages I (previtellogenic) and III (cortical rod) ovaries of the giant tiger shrimp (Penaeus monodon) were established. In all, 452 ESTs were unidirectionally sequenced. Sequence assembly generated 28 contigs and 201 singletons, 109 of which (48.0%) corresponding to known sequences previously deposited in GenBank. Several reproduction-related transcripts were identified. The full-length cDNA of anaphase promoting complex subunit 11 (PmAPC11; 600 bp with an ORF of 255 bp corresponding to a polypeptide of 84 amino acids) and selenoprotein Mprecursor (PmSePM; 904 bp with an ORF of 396 bp corresponding to a polypeptide of 131 amino acids) were characterized and reported for the first time in penaeid shrimp. Semiquantitative RT-PCR revealed that the expression levels of PmSePM and keratinocyte-associated protein 2 significantly diminished throughout ovarian development, whereas Ser/Thrcheckpoint kinase 1 (Chk1), DNA replication licensing factor mcm2 and egalitarian were down-regulated in mature ovaries of wild P. monodon (p < 0.05). Accordingly, the expression profiles of PmSePM and keratinocyte-associated protein 2 could be used as biomarkers for evaluating the degree of reproductive maturation in domesticated P. monodon.

Highlights

The giant tiger shrimp (Penaeus monodon) is one of the most economically important cultured species (BaileyBrock and Moss, 1992; Rosenberry, 2001)
Expression profiles of keratinocyte-associated protein 2, Ser/Thr checkpoint kinase 1, DNA replication licensing factor mcm2, P. monodon selenoprotein M precursor (PmSePM) and egalitarian during ovarian development of P. monodon broodstock were analyzed by way of semiquantitative RT-PCR
Homologues of thrombospondin (TSP; 39 ESTs accounting for 17.7% and ESTs accounting for 11.2% of sequenced clones) and peritrophin (39 ESTs, 17.7% and clones, 11.6%) were abundantly represented in both libraries similar to results from analyses of the conventional cDNA library of vitellogenic ovaries of P. monodon (79 and 87 clones accounting for 7.5 and 8.3% of clones sequenced, respectively; Preechaphol et al, 2007)

Summary

Introduction

The giant tiger shrimp (Penaeus monodon) is one of the most economically important cultured species (BaileyBrock and Moss, 1992; Rosenberry, 2001). The expression profiles of five reproduction-related genes during ovarian development in wild P. monodon broodstock were further examined using semiquantitative RT-PCR. ESTs representing P. monodon selenoprotein M precursor (PmSePM) and anaphase promoting complex subunit 11 (PmAPC11) were further sequenced from the reverse direction of the original cDNA clones by using a M13 forward primer.

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