STOCK Hand2^tm1.1Majh/AbfiJ

Stock number: 027727
Product name: Hand2 floxed (Hand2^loxp , Hand2^ploxp , Hand2^fl , Hand2^flox)
Research areas: Cancer Research, Cardiovascular Research, Cell Biology Research, Neurobiology Research, Research Tools

Description

The Hand2^fl allele has loxP sites flanking exon 1 (PGK-neo removed) of the Hand2 gene. These floxed mice are used to generate global or tissue-specific Hand2 deletion mice, which allow studies of this transcription factor in sympathetic nervous system morphogenesis and cell type-specific gene expression in sympathetic, enteric and adrenal chromaffin cells, cardiac morphogenesis and limb bud / branchial arch development. The absence of Hand2 expression may also be a useful biomarker for atypical hyperplasia, endometrioid carcinoma and epigenetic silencing.

Detailed description

The Hand2^fl allele has loxP sites flanking exon 1 of the Hand2 gene. Because the PGK-neo cassette is not present in this allele, heterozygotes (Hand2^fl/+) and homozygotes (Hand2^fl/fl) are viable and fertile with no alterations in Hand2 expression levels/patterns (in situ hybridization and qRT-PCR) compared to wildtype mice. Homozygotes exhibit no reported abnormal phenotype. Hand2 is a basic helix-loop-helix family transcription factor that functions in sympathetic nervous system morphogenesis and cell type-specific gene expression in sympathetic, enteric and adrenal chromaffin cells, cardiac morphogenesis and limb bud / branchial arch development. The absence of Hand2 expression may also be a useful biomarker for atypical hyperplasia, endometrioid carcinoma and epigenetic silencing.

When Hand2^fl mice are bred to Cre recombinase-expressing animals, the resulting offspring may be useful in generating global Hand2 knockout mice or tissue-specific Hand2 knockout mice. Specific examples are described below.

When bred to germline Cre-expressing mice, the resulting Hand2 global knockout homozygotes (Hand2^ko or Hand2^Delta) are embryonic lethal around embryonic day (E)10-12, with some surviving to birth but expiring thereafter.

Breeding Hand2^fl mice to Wnt1-Cre transgenic mice (Stock Nos. 003829 / 009107) results in deletion of Hand2 expression in neural crest cells. The resulting Hand2^cko homozygotes exhibit reduced neuronal precursor cell/neuroblast proliferation and neuronal numbers, abnormal facial/skull development and aglossia (absence of a tongue), with no gross cleft palate. Hand2^cko mice also have neurogenesis defects/decreased neurons in the developing enteric nervous system with few to no noradrenergic sympathetic ganglion neurons.

Deletion of Hand2 expression in neural precursor cells using Nestin-Cre transgenic mice (Stock Nos. 003771 / 019103) is lethal at around postnatal day (P)20, allowing analysis throughout development and the early neonatal period. Such mice have abnormal enteric nervous system development with loss of all nitergic neurons, decreased numbers of cholinergic neurons and functional aganglionosis.

Development

A targeting vector was designed by Dr. Marthe J. Howard (University of Toledo) to insert a loxP site upstream of exon 1, and a loxP-flanked PGK-neo cassette site downstream of exon 1 of the heart and neural crest derivatives expressed transcript 2 gene (Hand2) on chromosome 8. The construct was electroporated into 129S1/SvImJ-derived embryonic stem (ES) cells. Correctly targeted ES cells were were identified and injected into recipient blastocysts. The chimeric animals were bred to C57BL/6J mice to establish germline transmission. The Hand2^loxpneop allele was found to be a hypomorph, so the mice were then bred to Sycp1-Cre mice (undisclosed genetic background; Stock No. 003466) for germline removal of the PGK-neo cassette. The resulting offspring that retained the loxP-flanked exon 1 with the PGK-neo removed were selected and called Hand2^fl (also Hand2^loxp, Hand2^ploxp or Hand2^fl Hand2^flox). The Hand2^fl colony was maintained on a mixed C57BL/6;129S genetic background and the Cre transgene was removed. In 2015, Hand2^fl breeders were sent to Dr. Anthony B. Firulli (Indiana University School of Medicine). There, Hand2^fl mice were bred together to make homozygous. In 2015, Dr. Firulli sent Hand2^fl males on a mixed C57BL/6;129S genetic background (agouti coat colors) to The Jackson Laboratory Repository. Upon arrival, mice were backcrossed to C57BL/6J inbred mice (Stock No. 000664) to for at least two generations to establish our living Hand2^fl mouse colony (negative for neo and Cre). Later, sperm was cryopreserved.

Allele

Symbol: Hand2^tm1.1Majh
Name: targeted mutation 1.1, Marthe J Howard
MGI accession ID: MGI:3803647
Generation method: Targeted
Attributes: Conditional ready (e.g. floxed); No functional change
Synonyms: Hand2^ploxp
Marker symbol: Hand2
Marker name: heart and neural crest derivatives expressed 2
Marker synonyms: bHLHa26; dHAND; DHAND2; Ehand2; Hed; Thing2
Chromosome: 8
Strain of origin: 129S1/SvImJ
Molecular note: A loxP site was inserted upstream of exon 1 and a floxed neo cassette was inserted downstream of exon 1. Germ line, cre-mediated recombination was used to remove the neo cassette leaving exon 1 floxed.