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EPSA1 and VPF genes expression during embryonic and larval development period of Beluga, Huso huso
2016
Taheri Mirghaed, Ali
Background: The Endothelial PAS domain-containing protein 1 (EPSA1) is the key transcriptional regulator of hypoxic response and Vascular Permeability Factor (VPF) is an important growth factor for vascular development and angiogenesis. OBJECTIVES: In the present study, the levels of the EPSA1 coding gene and VPF transcripts were evaluated during Larval development of Beluga, Huso huso. METHODS: Samples at 12 developmental time-points including 1, 2, 4 days before hatch (eyed eggs), fresh hatched larvae (0), and larvae 1, 3, 6, 10, 15, 20, 25 and 50 days post-hatching were collected and stored in a −80 °C freezer until RNA extraction. Changes in EPSA1 and VPF mRNA expression were studied and differences in normalized mRNA expression levels among the different developmental stages of H. huso were analyzed by one-way analysis of variance (ANOVA). RESULTS: The transcripts of EPSA1 and VPF were detected in all developmental time-points of H. huso from embryos to fingerling fish. Our results revealed that the mRNA expression of EPSA1 and VPF was low during embryonic development and then upregulated significantly at the time of hatch and early larval time-points, whereas in the late larval development stages they started to decline. CONCLUSIONS: This study showed that there is an association between the EPSA1 and VPF mRNA expression during larval development of H. huso. The up regulation of EPSA1 and VPF transcripts at the time of hatch and during yolk sac fry development of H. huso is likely tied to the role of them in vasculogenesis and angiogenesis.
显示更多 [+] 显示较少 [-]Cloning and expression of Eimeria necatrix microneme5 gene in Escherichia coli
2016
Mayahi, Mansour | Jolodar, Abbas | Masaeli, Sharouz | Hamidinejat, Hosein | Seyfi Abad Shapouri, Masoud | Moori Bakhtiari, Naghme
Background: Coccidiosis caused by Eimeria necatrix has the most economic impact onpoultry production. Micronemal proteins in Eimeria necatrix are thoughtto be critical ligands determining host cell specificity at the time ofinvasion. OBJECTIVES: Isolation and purification of Eimeria necatrix oocysts from Khuzestan province of Iran was performed. AcDNA encoding microneme 5 (EnMIC5) protein was cloned and expressed asrecombinant protein before the evaluation of its immunogenicity by Westernblotting. METHODS: A primer pair was designed based on the publishednucleotide sequence of Eimeria necatrix LZ strain micronem5 gene. APartial cDNA sequence fragment of 758 bp coding for microneme 5 protein(EnMIC5) was amplified by semi- Nested RT-PCR. PCR products were cloned andexpressed in a Maltose Binding protein (MBP) containing expression vector(pMAL-c2x) in Escherichia coli. The cDNA which is encoded for 252 aminoacids shows high degree of conservation. It contains the adhesive plasmapre-kallikrein and seven hydrophilic motifs. RESULTS: The results of SDS-PAGErevealed that the recombinant protein with a molecular weight of 70 kDa wasover-expressed after induction with IPTG. Western blotting results revealedthat the expressed recombinant protein was reacted with sera of the chicksinfected with Eimeria necatrix. It was suggested that this proteinshould have a good immunogenicity and can be used for further studies. CONCLUSIONS: In conclusion, the high degree of sequence homology indicates that thisprotein is immunogenic and might be aninteresting vaccine target, and deserves further investigation
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