PCR-BASED ANALYSIS OF THE MURINE IMMUNOGLOBULIN HEAVY-CHAIN REPERTOIRE
PCR-BASED ANALYSIS OF THE MURINE IMMUNOGLOBULIN HEAVY-CHAIN REPERTOIRE
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DOI：
10.1016/0022-1759(95)00091-n
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发表时间：
1995-08-18
影响因子：
2.2
通讯作者：
KOURILSKY, P
中科院分区：
文献类型：
--
作者：
DELASSUS, S;GEY, A;KOURILSKY, P
主要通过研究属于所描述的 14 个家族中每个家族的基因在表达的免疫球蛋白分子方面的比例来分析免疫球蛋白重链库。尽管每个可变基因家族的比例在整个成年过程中以及在同一品系的不同小鼠中保持稳定，但关于激活的 B 细胞库中克隆代表性的信息很少。我们在这里描述了一种新方法，通过分离覆盖测序凝胶中免疫球蛋白重链基因的 V-H-D-J(H) 连接的聚合酶链式反应产物，从而允许使用给定的 J(H) 和给定 V-H 家族的成员之一区分不同的重排（根据其长度）来解决这个问题。使用这种方法，我们表明可以在来自 DNA 研究的 mRNA 水平以及潜在的库。我们还表明，这种方法可以检测诱导的免疫 B 细胞反应并研究新兴的显性特异性 B 细胞克隆。
The immunoglobulin heavy-chain repertoire has been mainly analysed by studying the proportion of genes belonging to each of the 14 described families, in terms of the expressed immunoglobulin molecules. Although the proportion of each variable gene family is kept stable throughout adult life and in different mice of the same strain, little information is available on the clonal representation in the repertoire of activated B cells. We describe here a new method that permits an approach to this question by separating the products of a polymerase chain reaction covering the V-H-D-J(H) junction of the immunoglobulin heavy-chain gene in a sequencing gel, thereby allowing discrimination of different rearrangements (according to their length) using a given J(H) and one of the member of a given V-H family.Using this method, we show that it is possible to obtain a precise overview of the repertoire of activated B cells, at the mRNA level, as well as the potential repertoire, from a study of the DNA. We also show that this approach permits the detection of an induced immune B cell response and studies of emerging dominant specific B cell clones.