Sox9 gene, a crucial member of the Sox gene family, is present in various organisms and involved in many physiological processes, especially in sex determination and gonad development. In this study, we reported a sox9 gene (designated as Spsox9) from Scylla paramamosain through analyzing published gonad transcriptome data. Meanwhile, the accuracy was validated by PCR technology, and the 3′ sequences were cloned with 3′ RACE technology. The full-length cDNA of Spsox9 is 2843 bp, consisting of a 243 bp 5′ UTR, an 1124 bp 3′ UTR, and a 1476 bp ORF encoding 491 amino acids. Furthermore, to better understand its conservation among crustacean species, the sox9 gene ortholog was identified in several other crustaceans species with their published transcriptome data, respectively. All of the Sox9 proteins identified in the current study had the common feature of Sox proteins (HMG domain) and were highly conserved among analyzed crustacean species. In all examined tissues, the Spsox9 was mainly expressed in the gonad (testis and ovary), eyestalk, and cerebral ganglion. During embryo development, Spsox9 was highly expressed in 5 pairs of appendages, 7 pairs of appendages, and eye-pigment formation stage. During ovary development, the expression level of Spsox9 remained stable in the first 4 stages (O1-O4) and decreased in the tertiary vitellogenesis (O5) stage. During testis development, the expression level of Spsox9 was highest in the spermatid stage (T2) and was significantly different from that in the spermatocyte stage (T1) and mature sperm stage (T3) (p < 0.05). In addition, Spsox9 exhibited a sex-biased expression pattern in T1 and O1. These present results indicated that the Spsox9 gene might play crucial roles in the gonad and embryo development of mud crab.

Sox9基因是Sox基因家族的重要成员，它存在于各种生物中，并参与许多生理过程，尤其是在性别确定和性腺发育中。在这项研究中，我们报道了SOX9基因（命名为Spsox9从）青蟹paramamosain通过分析公布的性腺转录数据。同时，通过PCR技术验证了准确性，并使用3'RACE技术克隆了3'序列。Spsox9的全长cDNA为2843 bp，由243 bp的5'UTR，1124 bp的3'UTR和1476 bp的ORF组成，编码491个氨基酸。此外，为了更好地了解其在甲壳类中的保守性，sox9该基因直系同源物分别在其他几个甲壳类物种中通过其已公开的转录组数据进行鉴定。在本研究中鉴定出的所有Sox9蛋白均具有Sox蛋白（HMG结构域）的共同特征，并且在所分析的甲壳类中高度保守。在所有检查过的组织中，Spsox9主要表达在性腺（睾丸和卵巢），眼球和脑神经节中。在胚胎发育过程中，Spsox9在5对附肢，7对附肢和眼色素形成阶段中高表达。在卵巢发育过程中，Spsox9的表达水平在前四个阶段（O1-O4）保持稳定，而在第三卵黄发生（O5）阶段下降。睾丸发育过程中的表达水平Spsox9在精子期（T2）最高，与精母细胞期（T1）和成熟精子期（T3）显着不同（p  <0.05）。此外，Spsox9在T1和O1中表现出性别偏向的表达模式。这些目前的结果表明，Spsox9基因可能在泥蟹的性腺和胚胎发育中起关键作用。