Utilization of a potentially universal downstream primer in the rapid identification and characterization of V lambda genes from two new human V lambda gene families.
Utilization of a potentially universal downstream primer in the rapid identification and characterization of V lambda genes from two new human V lambda gene families.
复制标题
利用潜在通用的下游引物快速鉴定和表征来自两个新的人类 V lambda 基因家族的 V lambda 基因。
DOI：
10.1111/j.1365-3083.1994.tb03345.x
复制
发表时间：
1994
影响因子：
3.7
通讯作者：
Chen,PP
中科院分区：
文献类型：
--
作者：
Deftos,M;Soto-Gil,R;Quan,M;Olee,T;Chen,PP
聚合酶链反应（PCR）极大地提高了克隆和鉴定大量基因的速度。然而，在鉴定和分析新的种系Ig - variable (V)基因家族方面，由于缺乏相关V基因下游侧翼区域的序列信息，该基因在V(D)J重排过程中被删除，因此其应用受到阻碍。为了避免这一问题，研究人员探索了一种可能性，即退化的下游引物可能与特定的上游引物结合使用，以克隆新的Vβ基因家族成员，因为人类对Vλ基因的了解远少于Vh和Vk基因。首先，通过与已建立的下游引物进行比较，考察了简并引物扩增已知Vλ1基因的可行性和特异性。结果是积极的。因此，利用退化引物克隆和表征了新近定义的Vλ8和Vλ9基因家族的种系Vλ基因。目前的策略可能有助于加快所有人类Vλ基因的鉴定和表征。此外，类似的策略可以应用于快速识别和表征已知或未知的Ig和T细胞受体V基因家族的新V基因。
Polymerase chain reaction (PCR) has increased dramatically the speed of cloning and characterizing numerous genes. However, its application to identifying and analysing new germline Ig‐variable (V) gene families has been hampered by the lack of sequence information in the downstream flanking regions of the concerned V genes, which are deleted during V(D)J rearrangements. To circumvent this problem, the possibility was explored that a degenerate downstream primer may be used in conjunction with a specific upstream primer, to clone members of new Vβ gene families, as much less is known about Vλ genes than Vh and Vk genes in humans. Firstly the feasibility and the specificity of a degenerate primer was examined by comparing it with an established downstream primer in amplifying known Vλ1 genes. The results were positive. Thus, the degenerate primer was used to clone and characterize germline Vλ genes of the recently defined Vλ8 and Vλ9 gene families. This current strategy may help speed up the identification and characterization of all human Vλ genes. Moreover, a similar strategy can be applied to identify and characterize rapidly new V genes of either known or unknown Ig and T‐cell receptor V gene families.