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利用靶向基因扩增和液滴数字 PCR 技术对 Tg(Adipoq-cre)1Evdr 小鼠脂联素启动子+ Cre 重组酶插入进行的特征描述。

Characterization of the adiponectin promoter + Cre recombinase insertion in the Tg(Adipoq-cre)1Evdr mouse by targeted locus amplification and droplet digital PCR.

机构信息

Section on Growth and Obesity and Division of Intramural Research, Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD), National Institutes of Health , Bethesda, MD, USA.

Section on Genetics and Endocrinology, Division of Intramural Research, Eunice Kennedy Shriver National Institute of Child Health and Human Development (NICHD), National Institutes of Health , Bethesda, MD, USA.

出版信息

Adipocyte. 2021 Dec;10(1):21-27. doi: 10.1080/21623945.2020.1861728.

Abstract

The Tg(Adipoq-cre)1Evdr mouse has become an important tool in adipose tissue biology. However, the exact genomic transgene integration site has not been established. Using Targeted Locus Amplification (TLA) we found the transgene had integrated on mouse chromosome 9 between exons 6 and 7 of . We detected transgene-transgene fusion; therefore, we used droplet digital polymerase chain reaction to identify copy number. In two separate experiments, we digested with BAMHI and with HindIII to separate potentially conjoined sequences. We found one copy of intact present in each experiment, indicating transgene-transgene fusion in other parts of the BAC that would not contribute to tissue-specific expression. copy number for Tg(Adipoq-cre)1Evdr mice can be potentially used to identify homozygous mice.

摘要

Tg(Adipoq-cre)1Evdr 小鼠已成为脂肪组织生物学的重要工具。然而,确切的基因组转基因整合位点尚未确定。使用靶向基因座扩增(TLA),我们发现转基因已整合到. 基因的第 6 号和第 7 号外显子之间的小鼠染色体 9 上。我们检测到转基因-转基因融合;因此,我们使用液滴数字聚合酶链反应来鉴定拷贝数。在两个独立的实验中,我们分别用 BamHI 和 HindIII 进行消化,以分离可能连接的. 序列。我们发现每个实验中都存在一个完整的 拷贝,表明 BAC 中的其他部位存在转基因-转基因融合,这不会导致组织特异性. 的表达。Tg(Adipoq-cre)1Evdr 小鼠的. 拷贝数可用于鉴定纯合子小鼠。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/c6fb/7781622/87197b940f50/KADI_A_1861728_F0001_OC.jpg

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