B7

acta2-optoβ2AR-IRES-lacZ Strain Information Sheet

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Strain Name:

Common Name: acta2-optoβ2AR-IRES-lacZ

CHROMusTM designation and lines available: R24:B7 Line 5

Jackson Stock number: NA

Development: The rhodopsin based effector, optoβ2AR which on activation generates the second messenger cAMP, was inserted at the ATG site of the acta2 gene in the BAC RP23-370F21 (CHORI) through homologous recombination. The resulting recombinant BAC was injected into the male pronucleus of fertilized oocytes which were then implanted into pseudo pregnant females. Resulting offspring were screened for the presence of the transgene (founders). Colonies were established from each founder and tested for expression.

Transgenic Numbers:

# pups born # founders # expressers
14 6 3

Description: The rhodopsin based effector, optoβ2AR is expressed under control of the acta2 promoter, directing expression to smooth muscle cells in blood vessels, airways of the lung and gut.  Expression may be present in other smooth muscle containing tissues which have not been examined.  An internal ribosome entry site (IRES) allows for co-expression of lacZ as a visual indicator of expression patterns.

Phenotypic Data:

qPCR analysis of optob2AR message in the bladder

qPCR analysis of optob2AR message in the bladder

 

lacZ staining of smooth muscle cells in the heart (line 6)

lacZ staining of smooth muscle cells in the heart (line 6)

 

Photo-activation of optoβ2AR. Blue light is flashed at t=15 seconds, inducing vasodilation in vascular smooth muscle.  See screenshots below.

 

b7-photo-activation-vasodilation-timelapse

 

Photo-activation in arterial myocytes showing vasodilation.

Photo-activation in arterial myocytes showing vasodilation.

b7-photoactivation-in-arterial-myocytes

Photo-activation of optob2AR increases Kv currents in arterial myocytes

 

b7-photoactivation-in-arterial-myocytes-2

Photo-activation of optob2AR increases Kv currents in arterial myocytes

 

 

 

 

 

 

 

Genotyping Protocol:

Primer B121 ATA GCT CTC AGC AAC CTG TTG GGT
Primer B72 TTC AAC ACG GTT TGG AGG CG
Expected size of product: 963bp

Cycling conditions:

94oC 3 minutes
Repeat 30 times
30 sec @ 94oC
30 sec @ 58oC
30 sec @ 72oC
72oC 5 minutes
10oC hold

Terms of Use:  Please contact chromus@cornell.edu for information on MTA and fees.

So as to appropriately acknowledge and credit the scientists who have produced the acta2-optoβ2AR-IRES-lacZ mouse strain, it is expected that you will include these individuals in any publication that is submitted prior to the initial publication of this line from the Kotlikoff laboratory.

Acknowledgement: Please inform us of any publications resulting from the use of this mouse and use the following statement to acknowledge their development:

The mouse strain acta2-optoβ2AR-IRES-lacZ was developed by CHROMusTM which is supported by the National Heart Lung Blood Institute of the National Institute of Health under award number R24HL120847.

 

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