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  • Novel method to rescue a lethal phenotype through integration of target gene onto the X-chromosome.

Novel method to rescue a lethal phenotype through integration of target gene onto the X-chromosome.

Scientific reports (2016-11-16)
Kazuya Sakata, Kimi Araki, Hiroyasu Nakano, Takashi Nishina, Sachiko Komazawa-Sakon, Shin Murai, Grace E Lee, Daisuke Hashimoto, Chigure Suzuki, Yasuo Uchiyama, Kenji Notohara, Anna S Gukovskaya, Ilya Gukovsky, Ken-Ichi Yamamura, Hideo Baba, Masaki Ohmuraya
摘要

The loss-of-function mutations of serine protease inhibitor, Kazal type 1 (SPINK1) gene are associated with human chronic pancreatitis, but the underlying mechanisms remain unknown. We previously reported that mice lacking Spink3, the murine homologue of human SPINK1, die perinatally due to massive pancreatic acinar cell death, precluding investigation of the effects of SPINK1 deficiency. To circumvent perinatal lethality, we have developed a novel method to integrate human SPINK1 gene on the X chromosome using Cre-loxP technology and thus generated transgenic mice termed "X-SPINK1". Consistent with the fact that one of the two X chromosomes is randomly inactivated, X-SPINK1 mice exhibit mosaic pattern of SPINK1 expression. Crossing of X-SPINK1 mice with Spink3+/- mice rescued perinatal lethality, but the resulting Spink3-/-;XXSPINK1 mice developed spontaneous pancreatitis characterized by chronic inflammation and fibrosis. The results show that mice lacking a gene essential for cell survival can be rescued by expressing this gene on the X chromosome. The Spink3-/-;XXSPINK1 mice, in which this method has been applied to partially restore SPINK1 function, present a novel genetic model of chronic pancreatitis.

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Sigma-Aldrich
胰蛋白酶 来源于牛胰腺, TPCK Treated, essentially salt-free, lyophilized powder, ≥10,000 BAEE units/mg protein
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抗 α-微管蛋白单克隆抗体 小鼠抗, ascites fluid, clone B-5-1-2
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DL-甘油醛-3-磷酸 溶液, 45-55 mg/mL in H2O
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Anti-Ras Antibody, clone RAS10, clone RAS10, Upstate®, from mouse
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甲基丙烯酸丙酮缩甘油酯, 50 wt. % in dichloromethane, contains ~280 ppm 4-tert-butylcatechol as inhibitor