三角帆蚌溶血磷脂酰胆碱酰基转移酶1HcLPCAT1基因功能分析及壳色性状相关SNP筛选
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张进盼(1996–),女,硕士研究生,研究方向为淡水贝类养殖和种苗工程.E-mail:jinpan_zhang@163.com

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S961

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国家自然科学基金项目(31872565); 财政部和农业农村部: 国家现代农业产业技术体系资助项目; 上海市优秀学术带头人计划资助项目(19XD1421500).


Functional analysis and SNP screening of lysophosphaticly lcholine acyltransferase 1 HcLPCAT1 gene and its association analysis with shell color traits in Hyriopsis cumingii
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    摘要:

    溶血磷脂酰胆碱酰基转移酶 1(LPCAT1)是一种重要的脂质代谢酶。为明确三角帆蚌(Hyriopsis cumingii) HcLPCAT1 基因在类胡萝卜素代谢中的功能, 探究该基因与三角帆蚌壳色的相关性, 本研究采用 RACE 技术克隆获得 HcLPCAT1 基因 cDNA 全长 1675 bp, ORF 区 1296 bp 编码 431 个氨基酸; 实时荧光定量分析发现 HcLPCAT1 基因在紫色三角帆蚌各组织中表达量均高于白色三角帆蚌相应组织, 且在肝胰腺、外套膜中的表达量差异显著 (P<0.05); 原位杂交检测到的阳性信号主要定位在外套膜的外褶、背膜区、腹膜区, 外褶与中褶连接处以及部分中褶; 紫色三角帆蚌补充投喂 β-胡萝卜素后, HcLPCAT1 基因在肝胰腺、中央膜、边缘膜各组织中表达量极显著上调 (P<0.01), 同时相应组织中总类胡萝卜素含量(TCC)极显著升高(P<0.01)。采用直接测序法鉴定出三角帆蚌 HcLPCAT1 基因 5 个 SNP 位点的基因型与内壳色存在显著相关性(P<0.05), 单倍型分析发现 H1、H2 两种单倍型为紫色三角帆蚌优势单倍型, H3、H5、H6 三种单倍型为白色三角帆蚌优势单倍型。本研究鉴定的 HcLPCAT1 基因可为解析三角帆蚌类胡萝卜素代谢和壳色形成的机制提供研究基础, 筛选的与内壳色相关 SNP 及单倍型可用于分子辅助育种。

    Abstract:

    Lysophosphaticly lcholine acyltransferase 1 (LPCAT1) is an important lipid metabolic enzyme. In order to clarify the function of HcLPCAT1 gene in carotenoid metabolism and the correlation of HcLPCAT1 gene with the shell nacre color in Hyriopsis cumingii. The cDNA full-length sequence of HcLPCAT1 gene in H. cumingii was cloned by RACE. Quantitative real-time PCR (qRT-PCR) and in situ hybridization (ISH) were used to analyze the expression pattern of HcLPCAT1 in different tissues. The effect of HcLPCAT1 on carotenoid metabolism was determined by supplementary feeding experiment. The direct sequencing method was used to screen the SNP sites related to inner shell color in the HcLPCAT1 gene. The results showed that the full-length cDNA of HcLPCAT1 was 1675bp with a 1296bp-long open reading frame (ORF) encoding 431 amino acids. HcLPCAT1 was expressed in the white and purple mussel tissues, with the expression levels being higher in the purple mussel than in the white mussel, and there was significant difference (P<0.05) between hepatopancreas and fringe mantle. The situ hybridization analysis showed that the positive signal of HcLPCAT1 located at the outer fold of mantle, dorsal mantle, ventral mantle, the joint of outer fold and middle fold of mantle and part of middle fold. After supplementation with β-carotene to purple mussel, the expression of HcLPCAT1 gene in hepatopancreas, middle mantle and fringe mantle increased significantly (P<0.01), and total carotenoids content (TCC) in these tissues also increased significantly (P<0.01). The genotypes of five SNP sites on the HcLPCAT1 gene were identified to be significantly correlated with the inner shell color parameters of H. cumingii. Haplotype analysis revealed that the H1 and H3 were the dominant haplotypes in the purple mussel, and the H3, H5 and H6 were the dominant haplotypes in the white mussel. The HcLPCAT1 gene identified in this study provides a molecular basis for analyzing the carotenoid metabolism and shell color formation of H. cumingii. The selected SNPs and haplotypes of HcLPCAT1 gene related to inner shell color can be used in molecular assisted breeding.

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张进盼,白志毅,张梦莹,颜玲,陆风辉,王贺.三角帆蚌溶血磷脂酰胆碱酰基转移酶1HcLPCAT1基因功能分析及壳色性状相关SNP筛选[J].中国水产科学,2021,28(11):1373-1384
ZHANG Jinpan, BAI Zhiyi, ZHANG Mengying, YAN Ling, LU Fenghui, WANG He. Functional analysis and SNP screening of lysophosphaticly lcholine acyltransferase 1 HcLPCAT1 gene and its association analysis with shell color traits in Hyriopsis cumingii[J]. Journal of Fishery Sciences of China,2021,28(11):1373-1384

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  • 在线发布日期: 2021-11-10
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