Functional analysis of gene LRRC8 causing agammaglobulinemia

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We previously isolated a novel gene, Leucine-rich repeat-containing 8 (LRRC8) which causes agammaglobulinemia in human. Transduction of mutated LRRC8 found in a patient into mouse hematopoietic cells lead to maturation block of B-cell progenitor at pro-B stage. In order to further elucidate the mechanism of congenital immunodeficiency, we tried making LRRC8-deficient mice. Targeting vector lacking the whole LRRC8 were made, and transfected into ES cells. Electroporation were performed twice: 64 samples at first time, 160 samples at second. Approximately 200 clones were picked up, and screened for positive clones. Positive clones were screened by Southern blot for right recombination, and one right clone was isolated. The ES clone (cell line) was injected into embryos to get germline chimeric mice, but birth of chmeric mice was unsuccessful.