Molecular Identification of Lobster Species Based on Cytochrome Oxidase Subunit I Gene characters.

<b>Background and Objective:</b> In the puerulus phase, which is not pigmented, identification of lobster species based on morphological characteristics is still difficult identity, so it is necessary to identify based on molecular characters. This study aimed to analyze the mitochondrial subunit I cytochrome oxidase (COI) gene characters of the puerulus of lobster species. <b>Materials and Methods:</b> The data can be useful for developing lobster seed identification methods based on DNA characteristics. Location of lobster sampling in Staring Bay, coastal waters of Moramo District, South Konawe Regency, Indonesia. The molecular characterization method is carried out in several stages, namely specific primer design, DNA preparation, PCR with specific primers, DNA sequencing and DNA sequence analysis. Characteristics of COI gene fragments were analyzed using BLAST analysis, restriction enzyme analysis and phylogenetic tree analysis. <b>Results:</b> The results showed that DNA was successfully isolated with a high level of purity. The results of the amplification of the COI gene fragment showed thick and firm bands and formed a single band measuring 751 bp with 249 amino acids. Based on the BLAST analysis shows that the COI gene fragment has 99% similarity with <i>Panulirus homarus</i>. Based restriction enzyme analysis shows that the site of recognition and restriction enzyme cutting position in the <i>Panulirus</i> COI gene fragment is the same as <i>Panulirus homarus</i> in Genebank, the <i>Ase</i>I enzyme in position 392 and <i>Psi </i>I in positions 47 and 106. <b>Conclusion:</b> Based on phylogenetic tree analysis, the COI gene fragment is in one group with <i>Panulirus homarus</i> and has a bootstrap value of 100% which shows that the nucleotide sequence is stable. The three analyzes show that the DNA source organism is the same species as <i>Panulirus homarus</i>.

Functional analysis of membrane-bound complement regulatory protein on T-cell immune response in ginbuna crucian carp.

Complements have long been considered to be a pivotal component in innate immunity. Recent researches, however, highlight novel roles of complements in T-cell-mediated adaptive immunity. Membrane-bound complement regulatory protein CD46, a costimulatory protein for T cells, is a key molecule for T-cell immunomodulation. Teleost CD46-like molecule, termed Tecrem, has been newly identified in common carp and shown to function as a complement regulator. However, it remains unclear whether Tecrem is involved in T-cell immune response. We investigated Tecrem function related to T-cell responses in ginbuna crucian carp. Ginbuna Tecrem (gTecrem) proteins were detected by immunoprecipitation using anti-common carp Tecrem monoclonal antibody (mAb) and were ubiquitously expressed on blood cells including CD8α(+) and CD4(+) lymphocytes. gTecrem expression on leucocyte surface was enhanced after stimulation with the T-cell mitogen, phytohaemagglutinin (PHA). Coculture with the anti-Tecrem mAb significantly inhibited the proliferative activity of PHA-stimulated peripheral blood lymphocytes, suggesting that cross-linking of Tecrems on T-cells interferes with a signal transduction pathway for T-cell activation. These findings indicate that Tecrem may act as a T-cell moderator and imply that the complement system in teleost, as well as mammals, plays an important role for linking adaptive and innate immunity.

Molecular characterization and expression analysis of three membrane-bound complement regulatory protein isoforms in the ginbuna crucian carp Carassius auratus langsdorfii.

Regulators of complement activation (RCA) play a role in protecting cells from excessive complement activation in humans. cDNA corresponding to three isoforms of teleost membrane-bound RCA protein (gTecrem) have been identified in the ginbuna crucian carp. gTecrem-1 consists of seven short consensus repeats (SCRs), whereas gTecrem-2 and gTecrem-3 have four SCRs. While gTecrem-1 possesses a tyrosine phosphorylation site in its cytoplasmic region, gTecrem-2 and gTecrem-3 lack the site. Tissue distribution analysis showed that gTecrem-1 and gTecrem-2 mRNAs were expressed in almost all tissues examined, whereas gTecrem-2 expression was not significantly detected in gill, liver, or intestine. Furthermore, analysis showed that gTecrem-1 was expressed in both peripheral blood leukocytes (PBLs) and erythrocytes and was also expressed in T cell subsets such as CD4(+), CD8(+) T cells, and IgM(+) B cells. gTecrem-2 expression was not detected in either PBLs or erythrocytes, whereas gTecrem-3 was expressed only in erythrocytes. These results suggested that gTecrem isoforms may serve different functional roles; gTecrem-1, which is expressed in T cells and possesses a tyrosine phosphorylation site, may act as a complement regulator and a cellular receptor in adaptive immunity.