Lin Y, Gross J K
Proc Natl Acad Sci U S A. 1981 May;78(5):2825-9. doi: 10.1073/pnas.78.5.2825.
Double-stranded cDNA was synthesized from partially purified winter flounder antifreeze mRNA and inserted into the endonuclease Pst I site of plasmid pBR322 by the poly(dG).poly(dC) homopolymer extension technique. The recombinant plasmids wee used to transform Escherichia coli. Clones containing antifreeze cDNA inserts were identified by the hybridization-selection technique. One of the inserts, 380 nucleotides in length, was digested with endonucleases Sau3AI and HinfI, which cleaved the insert into three fragments. The nucleotide sequences of these fragments were determined. The cDNA contains the entire coding sequence for a possible antifreeze peptide, including the leader sequence. The predicted amino acid sequence is similar to but not identical to one of the known sequences of antifreeze peptide. Within the cDNA are three segments of repeating sequences. The basic repeating sequence of 11 amino acids is maintained in the amino acid sequence coded by the cDNA and in the antifreeze peptide.
双链互补DNA是由部分纯化的冬鲽抗冻信使核糖核酸合成的，并通过聚（dG）·聚（dC）同聚物延伸技术插入到质粒pBR322的核酸内切酶Pst I位点。重组质粒用于转化大肠杆菌。含有抗冻互补DNA插入片段的克隆通过杂交选择技术进行鉴定。其中一个长度为380个核苷酸的插入片段用核酸内切酶Sau3AI和HinfI进行消化，这些酶将该插入片段切割成三个片段。测定了这些片段的核苷酸序列。该互补DNA包含一种可能的抗冻肽的完整编码序列，包括前导序列。预测的氨基酸序列与已知抗冻肽序列之一相似但不相同。在该互补DNA中有三个重复序列片段。由该互补DNA编码的氨基酸序列和抗冻肽中都保留了11个氨基酸的基本重复序列。
