Full-length transcriptome sequencing integrated with RNA-Seq reads identify pigment genes in goldfish (Carassius auratus) (III)

The goldfish (carassius auratus) is a valuable ornamental fish with the most diverse pheotypes, which is originally demoesticated from wild grey cruician carp in China over 1,000 yeas ago. Red color skin is the first trait fixed in goldfish and distinguished from its ancestor. However, the genomic resources about the speices is limited when performing the study, which heavely hampers our understanding of the genetic basis of the diverse phenotypes. To quickly provide a large amount of genomic resources and decipher the possible mechanism underlying diverse color skin in goldfish, We performed a large scale of transcriptome sequencing on 13 tissues and 4 typical color skins through combinng PacBio long-read sequencing and Illumina short-read sequencing. A full-length transcriptome with 137,674 transcripts was generated with mean length and N50 length of 2,956 bp and 4,017 bp, respectively. A total of 108,122 (78.53%) novel isoforms from known genes and 17,622 novel genes were identified compared to the annotation of recently published goldfish reference genome. Moreover, 59,014 alternative splicing events were present in 15,380 genes. A total of 162 differentially expressed genes (DEGs) were identified among the four different color skins, which were mainly involved in the pathways of Melanogenesis, Tyrosine metabolism, Riboflavin metabolism, Folate biosynthesis, and alpha-Linolenic acid metabolism. Fourteen DEGs may have function in pigmentation, including Melanophilin genes, solute carrier family 2 member 11 and solute carrier family 2 member 9. In conclusion, the genomic resources provided in study significantly improved the gene annotation of the published referecne genome and enhanced the understanding of the goldfish pigmentation pathways and will finally facilitate future studies on genetic basis of goldfish diverse phenotypes.

金鱼（Carassius auratus）是一种表型最为多样的珍贵观赏鱼类，其祖先为中国1000余年前驯化的野生灰色鲫。红色皮肤是金鱼首个固定下来、区别于其祖先的性状。然而当前该物种的基因组资源较为匮乏，严重阻碍了我们对其多样表型遗传基础的解析。为快速获取大量基因组资源并解析金鱼多样体色形成的潜在机制，本研究结合PacBio长读长测序（PacBio long-read sequencing）与Illumina短读长测序（Illumina short-read sequencing）技术，对13种组织及4种典型体色样本开展了大规模转录组测序（transcriptome sequencing）。本研究最终获得了包含137674条转录本的全长转录组（full-length transcriptome），其平均长度与N50长度分别为2956 bp与4017 bp。与近期发表的金鱼参考基因组注释信息相比，共鉴定得到108122条已知基因的新可变剪接异构体（占比78.53%）以及17622个新基因。此外，在15380个基因中共发现59014个可变剪接事件（alternative splicing events）。在4种不同体色样本间共鉴定到162个差异表达基因（differentially expressed genes, DEGs），这些基因主要富集于黑素生成（Melanogenesis）、酪氨酸代谢、核黄素代谢、叶酸生物合成以及α-亚麻酸代谢通路。其中14个差异表达基因可能参与色素沉着过程，包括黑素亲和素基因（Melanophilin genes）、溶质载体家族2成员11（solute carrier family 2 member 11）与溶质载体家族2成员9（solute carrier family 2 member 9）。综上，本研究提供的基因组资源显著优化了已发表参考基因组的基因注释，加深了我们对金鱼色素沉着通路的理解，将为后续解析金鱼多样表型的遗传基础提供重要支撑。