Hiraiwa A, Seyfried C E, Nepom G T, Milner E C
Virginia Mason Research Center, Seattle, WA 98101.
Immunogenetics. 1989;29(3):186-90. doi: 10.1007/BF00373644.
HLA class II allelic variants within the DQw3-related family of genes carry distinct allo-specificities and have been implicated in specific HLA-disease associations, such as insulin-dependent diabetes mellitus. To investigate the nucleotide variations which characterize DQw3 genes, we applied a novel cDNA cloning strategy that uses a single-stranded vector/primer system to facilitate DNA sequencing of allelically variable gene families. Using a DQB-specific primer sequence and M13 bacteriophage as the cloning vector, direct cloning and sequencing of multiple DQB genes was performed without the need for second strand synthesis or for subcloning. Sequence analysis from eight lymphoblastoid cell lines selected to represent different ethnic backgrounds revealed three DQw3-related DQB genes, DQB3.1, 3.2, and 3.3, corresponding to the newly designated HLA-DQw7, w8, and w9 specificities, respectively. An unusual Pro-Pro couplet at codons 55-56 is characteristic of all DQw3-positive sequences and may be contributing to the broad DQw3 allospecificity. Comparisons among ethnically disparate DQw3-related sequences showed no additional expressed or silent nucleotide substitutions among these DQB alleles. Thus, polymorphism within the DQw3 family of genes appears to be extremely limited, with a paucity of nucleotide variations accumulated by evolutionary distance.
DQw3相关基因家族中的HLA - II类等位基因变体具有独特的同种特异性，并与特定的HLA - 疾病关联有关，如胰岛素依赖型糖尿病。为了研究表征DQw3基因的核苷酸变异，我们应用了一种新颖的cDNA克隆策略，该策略使用单链载体/引物系统来促进等位基因可变基因家族的DNA测序。使用DQB特异性引物序列和M13噬菌体作为克隆载体，无需进行第二链合成或亚克隆即可直接克隆和测序多个DQB基因。对选自不同种族背景的八个淋巴母细胞系进行序列分析，发现了三个与DQw3相关的DQB基因，即DQB3.1、3.2和3.3，分别对应于新指定的HLA - DQw7、w8和w9特异性。密码子55 - 56处不寻常的脯氨酸 - 脯氨酸二联体是所有DQw3阳性序列的特征，可能有助于广泛的DQw3同种特异性。不同种族的DQw3相关序列之间的比较显示，这些DQB等位基因之间没有额外的表达或沉默核苷酸替换。因此，DQw3基因家族内的多态性似乎极其有限，由于进化距离积累的核苷酸变异很少。
