A novel strategy for conditional gene knockout based on ΦC31 integrase and Gal4/UAS system inDrosophila

The gut microbiota is very important in the initiation, progression, and dissemination of cancer, and the regulation of microbiota has been employed as a novel strategy to enhance the effect of immunotherapy. Adiponectin (APN), an adipocyte-derived hormone, plays a vital role in regulating the immune response of innate immune cells. The deficiency of APN inhibits rhabdomyosarcoma growth. However, whether this function is associated with regulating gut microbiota remains unknown. To investigate, we performed 16S ribosomal RNA (rRNA) gene sequencing on the fecal microbiome of APN gene knockout mice to determine whether APN deletion affects the gut microbiota. We found APN deficiency alters gut microbial functions involved in metabolism, genetic information processing, and cellular processes. In addition, a decreased abundance of Bacteroides and an increased abundance of Prevotella and Helicobacter were observed in rhabdomyosarcoma-bearing APN knockout mice; these bacteria were associated with the inhibition of rhabdomyosarcoma growth. These findings suggest that gut microbiota may be a potential target of APN deficiency against rhabdomyosarcoma.

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Efficient generation of zebrafish maternal-zygotic mutants through transplantation of ectopically induced and Cas9/gRNA targeted PGCs

10.1101/693853 ◽

2019 ◽

Author(s):

Fenghua Zhang ◽

Xianmei Li ◽

Mudan He ◽

Ding Ye ◽

Feng Xiong ◽

...

Keyword(s):

High Efficiency ◽

Essential Gene ◽

Gene Knockout ◽

Primordial Germ Cells ◽

Survival Rates ◽

Head Development ◽

Embryonic Lethal ◽

Novel Strategy ◽

Genome Activation ◽

Lethal Genes

ABSTRACTThe CRISPR/Cas9 technology has been widely utilized for knocking out genes involved in various biological processes in zebrafish. Despite this technology is efficient for generating different mutations, one of the main drawbacks is low survival rates during embryogenesis when knocking out some embryonic lethal genes. To overcome this problem, we developed a novel strategy using a combination of CRISPR/Cas9 mediated gene knockout with primordial germ cells (PGCs) transplantation to facilitate and speed up the process of zebrafish mutant generation, particularly for embryonic lethal genes. First, we optimized the procedure for gRNA targeted PGCs transplantation (PGCT), by increasing the efficiencies of genome mutation in PGCs and induction of PGCs fates in donor embryos for PGCT. Second, the combined CRISPR/Cas9 with PGCT was utilized for generation of maternal zygotic (MZ) mutants of tcf7l1a (essential gene for head development), pou5f3 (essential gene for zygotic genome activation) and chd (essential gene for dorsal development) at F1 generation with high efficiency. Finally, we revealed some novel phenotypes in the maternal zygotic mutant of tcf7l1a and chd, while MZtcf7l1a showed elevated neural crest development, and MZchd have stronger ventralization than its zygotic counterparts. Therefore, this study presents an efficient and powerful method for generating MZ mutants of embryonic lethal genes in zebrafish.

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Conditional Gene Knockout

Gene Biotechnology ◽

10.1201/b10919-27 ◽

2016 ◽

pp. 513-530

Keyword(s):

Gene Knockout ◽

Conditional Gene Knockout

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Essential Role of Chicken Ovalbumin Upstream Promoter-Transcription Factor II in Insulin Secretion and Insulin Sensitivity Revealed by Conditional Gene Knockout

Diabetes ◽

10.2337/diabetes.54.5.1357 ◽

2005 ◽

Vol 54 (5) ◽

pp. 1357-1363 ◽

Cited By ~ 28

Author(s):

P. Bardoux ◽

P. Zhang ◽

D. Flamez ◽

A. Perilhou ◽

T. A. Lavin ◽

...

Keyword(s):

Transcription Factor ◽

Insulin Secretion ◽

Insulin Sensitivity ◽

Essential Role ◽

Gene Knockout ◽

Factor Ii ◽

Upstream Promoter ◽

Conditional Gene Knockout ◽

Chicken Ovalbumin

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Conditional Gene Knockout Using Cre Recombinase

Molecular Biotechnology ◽

10.1385/mb:17:3:269 ◽

2001 ◽

Vol 17 (3) ◽

pp. 269-276 ◽

Cited By ~ 46

Author(s):

Yunzheng Le ◽

Brian Sauer

Keyword(s):

Gene Knockout ◽

Cre Recombinase ◽

Conditional Gene Knockout

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Identification of Jmjd3 as an Essential Epigenetic Regulator of Hox Gene Temporal Collinear Activation for Body Axial Patterning in Mice

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.642931 ◽

2021 ◽

Vol 9 ◽

Author(s):

Feng Zhang ◽

Xiong Zhao ◽

Runmin Jiang ◽

Yuying Wang ◽

Xinli Wang ◽

...

Keyword(s):

Gene Knockout ◽

Underlying Mechanism ◽

Axial Patterning ◽

Hox Gene ◽

Skeletal Patterning ◽

Epigenetic Regulator ◽

Gene Knockout Mice ◽

Demethylase Activity ◽

A Subunit ◽

Conditional Gene Knockout

Body axial patterning develops via a rostral-to-caudal sequence and relies on the temporal colinear activation of Hox genes. However, the underlying mechanism of Hox gene temporal colinear activation remains largely elusive. Here, with small-molecule inhibitors and conditional gene knockout mice, we identified Jmjd3, a subunit of TrxG, as an essential regulator of temporal colinear activation of Hox genes with its H3K27me3 demethylase activity. We demonstrated that Jmjd3 not only initiates but also maintains the temporal collinear expression of Hox genes. However, we detected no antagonistic roles between Jmjd3 and Ezh2, a core subunit of PcG repressive complex 2, during the processes of axial skeletal patterning. Our findings provide new insights into the regulation of Hox gene temporal collinear activation for body axial patterning in mice.

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