猪D-氨基酸氧化酶：通过基因组和cDNA克隆的特性鉴定来确定mRNA核苷酸序列

Porcine D-amino acid oxidase: determination of the mRNA nucleotide sequence by the characterization of genomic and cDNA clones.

作者信息

Jacobs P, Brockly F, Massaer M, Loriau R, Guillaume J P, Ciccarelli E, Heinderyckx M, Cravador A, Biemans R, van Elsen A

机构信息

Service for Applied Genetics, University of Brussels, Nivelles, Belgium.

出版信息

Gene. 1987;59(1):55-61. doi: 10.1016/0378-1119(87)90266-6.

DOI:10.1016/0378-1119(87)90266-6

PMID:2893757

Abstract

Oligodeoxynucleotide probes derived from the published amino acid (aa) sequence for D-amino acid oxidase (DAO) [Ronchi et al. J. Biol. Chem. 259 (1982) 8824-8834] were used to screen cDNA libraries made from porcine kidney cortex and liver. Whereas no clones were obtained from kidney mRNAs, 20 independent ones were isolated from the liver library. Surprisingly, all of them carried only partial cDNAs for DAO starting around aa 100 in the coding sequence and extending for up to 250 bp in the 3'-noncoding sequence. One of these clones, pULB9103, was used to screen a porcine genomic library and allowed the isolation of DAO gene clone phULB001. Four exons encoding aa 1-151 were identified and sequenced, as well as the relevant exon-intron junctions. The mRNA sequence coding for DAO has been reconstituted from the genomic and cDNA sequences; its analysis by computer did not reveal any significant secondary structure, or particular feature, which could explain the failure to obtain full-length cDNAs.

摘要

从已发表的D - 氨基酸氧化酶（DAO）氨基酸（aa）序列[Ronchi等人，《生物化学杂志》259 (1982) 8824 - 8834]衍生的寡脱氧核苷酸探针，用于筛选由猪肾皮质和肝脏构建的cDNA文库。虽然从肾mRNA未获得克隆，但从肝脏文库中分离出了20个独立克隆。令人惊讶的是，它们所有都仅携带DAO的部分cDNA，起始于编码序列中约第100个氨基酸左右，并在3' - 非编码序列中延伸多达250 bp。其中一个克隆pULB9103用于筛选猪基因组文库，并使得DAO基因克隆phULB001得以分离。鉴定并测序了编码第1 - 151个氨基酸的四个外显子以及相关的外显子 - 内含子连接。编码DAO的mRNA序列已从基因组和cDNA序列中重建；通过计算机分析未发现任何可解释未能获得全长cDNA的显著二级结构或特殊特征。