Orlandi R, Güssow D H, Jones P T, Winter G
Division of Experimental Oncology E, Istituto Nazionale per lo Studio e la Cura dei Tumori, Milan, Italy.
Proc Natl Acad Sci U S A. 1989 May;86(10):3833-7. doi: 10.1073/pnas.86.10.3833.
We have designed a set of oligonucleotide primers to amplify the cDNA of mouse immunoglobulin heavy and light chain variable domains by the polymerase chain reaction. The primers incorporate restriction sites that allow the cDNA of the variable domains to be force-cloned for sequencing and expression. Here we have applied the technique to clone and sequence the variable domains of five hybridoma antibodies and to express a mouse-human chimeric antibody that binds to the human mammary carcinoma line MCF-7. The technique should also lead to the cloning of antigen-binding specificities directly from immunoglobulin genes.
我们设计了一组寡核苷酸引物，通过聚合酶链反应扩增小鼠免疫球蛋白重链和轻链可变区的cDNA。这些引物含有限制性酶切位点，可将可变区的cDNA进行强制克隆以便测序和表达。在此，我们已应用该技术克隆并测序了五种杂交瘤抗体的可变区，并表达了一种与人类乳腺癌细胞系MCF-7结合的小鼠-人嵌合抗体。该技术还应能直接从免疫球蛋白基因克隆抗原结合特异性。
