C57BL/6-Tg(RP23-352G18)1Coppa/J

Stock number: 028361
Product name: Sirt6BAC
Research areas: Cardiovascular Research, Diabetes and Obesity Research, Metabolism Research, Research Tools

Description

This Sirt6BAC transgenic line exhibits eutopic and physiological overexpression of mouse SIRT6, resulting in enhanced insulin sensitivity in skeletal muscle and liver, as well as protection against aberrant glucose homeostasis and diet-induced type 2 diabetes mellitus.

Detailed description

Sirtuins are nicotinamide adenine dinucleotide-dependent enzymes exerting post-translational modifications of their target proteins, and are potential therapeutic targets for improving metabolic imbalances caused by high-calorie-diet. SIRT6, one of the seven mammalian sirtuins, is a lysine-deacetylase and mono-ADP-ribosyl-transferase with pleiotropic effects.

Mice that are hemizygous for the RP23-352G18 BAC transgene (Sirt6BAC line 1) have moderate, physiological overexpression of mouse SIRT6 (two-to-four fold greater than wildtype littermates). Sirt6BAC line 1 mice have normal body weight and adiposity, but increased circulating levels of polyamine spermidine. Sirt6 overexpression enhances insulin sensitivity in skeletal muscle and liver, and also imparts protection from high-calorie-diet-induced hyperglycemia and glucose intolerance. Specifically, compared to wildtype mice, Sirt6BAC line 1 has increased insulin-stimulated glucose uptake in both gastrocnemius and soleus muscle (but not in brain, interscapular brown adipose or epididymal adipose tissue), and increased insulin-induced p-AKT/AKT ratio in gastrocnemius muscle.

Hemizygous mice are viable and fertile. The mRNA expression level of the other genes on the BAC are described in the publication. To date (November 2015), the transgene insertion site and copy number is not known, and it has not been attempted to make homozygous mice.

Development

The Sirt6BAC transgenic mouse line was created by Dr. Roberto Coppari (University of Geneva). First, the ~185.7 kbp C57BL/6J mouse bacterial artificial chromosome (BAC) RP23-352G18 was obtained; containing the entire sirtuin 6 locus (Sirt6) as well as ~109 kbp of centromeric flanking sequences (including a portion of the Gna15 locus, several complete loci [Gna11, Aes, Tle2, Tle6, BC025920, Ankrd24] and predicted genes [Gm31878, Gm31936, Gm32024]) and ~70 kbp of telomeric flanking sequences (including the predicted gene Gm10778). The RP23-352G18 BAC DNA was modified only by insertion of an ampicillin resistance cassette, and the ~186.6 kbp construct was microinjected into the pronuclei of fertilized C57BL/6J embryos. Founder animals were bred to C57BL/6J inbred mice for germline transmission, establishing three founder lines. Sirt6BAC founder line 1 was identified with two-to-four fold greater expression of SIRT6 compared to wildtype littermates. The donating investigator reports that Sirt6BAC transgenic mice from this founder line were maintained by breeding to wildtype (non-transgenic) siblings each generation for more than five generations, and then sent to Dr. Raul Mostoslavsky (Massachusetts General Hospital, Harvard Medical School) in 2014. There, the mice were bred once to C57BL/6NCrl females, and then maintained by breeding to wildtype (non-transgenic) siblings prior to sending to The Jackson Laboratory Repository in 2015 (see SNP results below). Upon arrival, sperm was cryopreserved. To establish our living colony, an aliquot of the frozen sperm was used to fertilize oocytes from C57BL/6J inbred females (Stock No. 000664).

In 2016, a 32 SNP (single nucleotide polymorphism) panel analysis, with 27 markers covering all 19 chromosomes and the X chromosome, as well as 5 markers that distinguish between the C57BL/6J and C57BL/6N substrains, was performed on the first generation rederived living colony at The Jackson Laboratory Repository. All the 27 markers throughout the genome suggested a C57BL/6 genetic background. 5 of 5 markers that determine C57BL/6J from C57BL/6N were found to be segregating in some mice (chromosomes 8 [~15.2 Mbp], 11 [~4.4 Mbp], 13 [~41.0 Mbp] and 15 [~57.6 Mbp]. As expected, these data suggest that the mice sent to The Jackson Laboratory Repository were on a mixed C57BL/6N;C57BL/6J genetic background.

Allele

Symbol: Tg(RP23-352G18)1Coppa
Name: transgene insertion 1, Roberto Coppari
MGI accession ID: MGI:5696226
Generation method: Transgenic
Attributes: Inserted expressed sequence
Marker symbol: Tg(RP23-352G18)1Coppa
Marker name: transgene insertion 1, Roberto Coppari
Chromosome: UN
Strain of origin: C57BL/6J
Expressed genes: Sirt6,sirtuin 6,mouse, laboratory
Site of expression: Sirt6 along with other alleles in the BAC insertion are over-expressed in brain, gastrocnemius, and liver.
Molecular note: The transgenic construct was made using C57BL/6J mouse bacterial artificial chromosome (BAC) RP23-352G18 containing the entire sirtuin 6 locus (Sirt6) as well as 109 kbp of centromeric flanking sequences (including a portion of the Gna15 locus, several complete loci [Gna11, Aes, Tle2, Tle6, BC025920, Ankrd24] and predicted genes [Gm31878, Gm31936, Gm32024]) and 70 kbp of telomeric flanking sequences (including the predicted gene Gm10778). The RP23-352G18 BAC DNA was modified only by insertion of an ampicillin resistance cassette. Founder line 1 has a 2-4 fold greater expression of Sirt6 as compared to wild-type.