沙虾眼柄中一种推测的脱壳抑制激素cDNA的克隆。

P L Gu, S M Chan
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引用次数: 0

摘要

利用日本对虾(Penaeus japonicus)神经肽Pej-SGP-IV的氨基酸序列设计了退化引物。利用沙虾眼柄互补DNA进行逆转录聚合酶链反应(RT-PCR)。克隆了部分编码Pej-SG-IV神经肽同源蛋白的cDNA。以部分cDNA为探针,筛选眼柄cDNA文库。分离到多个与部分cDNA核苷酸序列相同的cDNA克隆。cDNA全长957 bp,开放阅读框由315 bp的编码序列组成。该神经肽的推导氨基酸由77个氨基酸组成,前面是28个氨基酸的信号肽。由于推导出的虾神经肽的氨基酸序列与P. japonicus的Pej-SGP-IV(抑制脱皮)和其他甲壳类动物的脱皮抑制激素(MIHs)高度同源,因此暂时将虾神经肽命名为MeMIH。Northern blot和RT-PCR分析显示,MeMIH在虾眼柄和脑的蜕皮后、蜕皮间和蜕皮前阶段均有表达。此外,在新发育的幼虫的神经组织中也可以检测到RNA信息。然而,MeMIH在肌肉、游泳腿和肝胰腺中没有发现。基因组Southern blot分析和聚合酶链反应(PCR)扩增结果表明,该虾基因组中存在一个MIH基因拷贝。推定的虾类MIH基因的结构组织与feriatus蟹类相似。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
Cloning of a cDNA encoding a putative molt-inhibiting hormone from the eyestalk of the sand shrimp Metapenaeus ensis.

Degenerate primers were designed from the amino acid sequence of the neuropeptide Pej-SGP-IV of the shrimp Penaeus japonicus. Reverse transcriptase-polymerase chain reaction (RT-PCR) was performed using eyestalk complementary DNA of the sand shrimp Metapenaeus ensis. A partial cDNA that codes for a protein homologous to the neuropeptide Pej-SG-IV was cloned. The partial cDNA was used as a probe to screen the eyestalk cDNA library. Several cDNA clones with nucleotide sequence identical to the partial cDNA were isolated. The largest cDNA is 957 bp with an open reading frame consisting of a coding sequence 315 bp in length. The deduced amino acid of the neuropeptide consists of 77 amino acids and is preceded by a signal peptide of 28 amino acids. Because the deduced amino acid sequence of the shrimp cDNA is highly homologous to the Pej-SGP-IV of P. japonicus (which is molt inhibiting) and to other crustaceans' molt-inhibiting hormones (MIHs), the shrimp neuropeptide is tentatively called MeMIH. Northern blot analysis and RT-PCR showed that MeMIH is expressed in the postmolt, intermolt, and premolt stages of the shrimp eyestalks and the brain. Moreover, RNA message can also be detected in the nervous tissues of newly developed larvae. MeMIH is, however, not found in the muscle, swimming leg, and hepatopancreas. Results from genomic Southern blot analysis and amplification of the shrimp genomic DNA by polymerase chain reaction (PCR) suggest that a single copy of the MIH gene is present in the genome. The structural organization of the gene for the shrimp putative MIH is similar to that of the crab Charybdis feriatus.

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