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Moult-Inhibiting Hormone from the Swimming Crab, Portunus Trituberculatus (Miers, 1876): PCR Cloning, Tissue Distribution, and Expression of Recombinant Protein in Escherichia Coli (Migula, 1895)

Moult-Inhibiting Hormone from the Swimming Crab, Portunus Trituberculatus (Miers, 1876): PCR... <jats:sec><jats:title>Abstract</jats:title><jats:p>In the present study, a genomic DNA of MIH (GenBank: #EU869539) was cloned from the swimming crab, Portunus trituberculatus (Miers, 1876). The genome DNA, consisting of 2865 bp, is comprised of three exons interrupted by two introns. Multiple sequence alignments revealed that in the 5 upstream region of MIH, sequences with high similarity to arthropod initiator, TATA box, CREB (cyclic AMP response element binding) protein were the common structure. The signal peptide in the genomic DNA was encoded by exon1 and exon2, which was interrupted by 242 bp-intron (intron1), located between gln12 and arg13. The mature peptide was encoded by exon2 and exon3, which was interrupted by 313 bp-intron (intron2), at the position between 2nd and 3rd nucleotides of the codon encoding arg41. Pot-MIH was expressed only in the eyestalk ganglia, ovaries, testes, posterior spermatic duct, bristle in ejaculatory duct, cranial ganglia, and thoracic ganglia, as determined in various tissues by semi-quantitative RT-PCR. A cDNA encoding the mature peptide was used to express recombinant MIH (rMIH) using the Escherichia coli (Migula, 1895) expression system. Two constructs were designed to yield either a mature MIH fusion protein with and without histidine (His) tag at the carboxyl terminus. The rMIH protein was detected by SDS-PAGE and Western blot analysis, indicating that the antibody prepared by two rMIH proteins has high specificity.</jats:p> </jats:sec> http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Crustaceana Brill

Moult-Inhibiting Hormone from the Swimming Crab, Portunus Trituberculatus (Miers, 1876): PCR Cloning, Tissue Distribution, and Expression of Recombinant Protein in Escherichia Coli (Migula, 1895)

Zhu, D. F.; Hu, Z. H.; Shen, J. M.
Crustaceana , Volume 84 (12-13): 1481 – Jan 1, 2011
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Publisher
Brill
Copyright
© 2011 Koninklijke Brill NV, Leiden, The Netherlands
ISSN
0011-216x
eISSN
1568-5403
DOI
10.1163/156854011X607051
Publisher site
See Article on Publisher Site

Abstract

<jats:sec><jats:title>Abstract</jats:title><jats:p>In the present study, a genomic DNA of MIH (GenBank: #EU869539) was cloned from the swimming crab, Portunus trituberculatus (Miers, 1876). The genome DNA, consisting of 2865 bp, is comprised of three exons interrupted by two introns. Multiple sequence alignments revealed that in the 5 upstream region of MIH, sequences with high similarity to arthropod initiator, TATA box, CREB (cyclic AMP response element binding) protein were the common structure. The signal peptide in the genomic DNA was encoded by exon1 and exon2, which was interrupted by 242 bp-intron (intron1), located between gln12 and arg13. The mature peptide was encoded by exon2 and exon3, which was interrupted by 313 bp-intron (intron2), at the position between 2nd and 3rd nucleotides of the codon encoding arg41. Pot-MIH was expressed only in the eyestalk ganglia, ovaries, testes, posterior spermatic duct, bristle in ejaculatory duct, cranial ganglia, and thoracic ganglia, as determined in various tissues by semi-quantitative RT-PCR. A cDNA encoding the mature peptide was used to express recombinant MIH (rMIH) using the Escherichia coli (Migula, 1895) expression system. Two constructs were designed to yield either a mature MIH fusion protein with and without histidine (His) tag at the carboxyl terminus. The rMIH protein was detected by SDS-PAGE and Western blot analysis, indicating that the antibody prepared by two rMIH proteins has high specificity.</jats:p> </jats:sec>

Journal

Crustaceana – Brill

Published: Jan 1, 2011

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