Experimental evaluation does not reveal a direct effect of microRNA from the callipyge locus on DLK1 expression.

Experimental evaluation does not reveal a direct effect of microRNA from the callipyge locus on DLK1 expression.

BMC genomics (2014-11-02)

Huijun Cheng, Xuewen Xu, Tracy Hadfield, Noelle Cockett, Carole Charlier, Michel Georges, Haruko Takeda

ABSTRACT

Polar overdominance at the ovine callipyge (CLPG) locus involves the post-transcriptional trans-inhibition of DLK1 in skeletal muscle of CLPG/CLPG sheep. The abundant maternally expressed microRNAs (miRNAs) mapping to the imprinted DLK1-GTL2 domain are prime candidate mediators of this trans-effect. We have tested the affinity of 121 miRNAs processed from this locus for DLK1 by co-transfecting COS1 cells with a vector expressing the full-length ovine DLK1 with corresponding mimic miRNAs. None of the tested miRNAs was able to down regulate DLK1 to the extent observed in vivo. This suggests that other factors, with or without these miRNAs, are involved in mediating the observed trans-effect.

MATERIALS

Product Number

Brand

Product Description

PHG0004

SAFC

BIS-TRIS

RES1161B-A7

SAFC

BIS-TRIS

14879

Sigma-Aldrich

BIS-TRIS, BioUltra, ≥99.0% (NT)

69947

Sigma-Aldrich

MOPS, BioUltra, for molecular biology, ≥99.5% (titration)

M1254

Sigma-Aldrich

MOPS, ≥99.5% (titration)

M5162

Sigma-Aldrich

MOPS, BioXtra, ≥99.5% (titration)

M3183

Sigma-Aldrich

MOPS, BioPerformance Certified, suitable for cell culture, ≥99.5% (titration)

B9754

Sigma-Aldrich

BIS-TRIS, ≥98.0% (titration)

B4429

Sigma-Aldrich

BIS-TRIS, BioPerformance Certified, suitable for cell culture, suitable for insect cell culture, ≥98.0%

B7535

Sigma-Aldrich

BIS-TRIS, BioXtra, ≥98.0% (titration)

PHG0007

SAFC

MOPS

RDD003

Sigma-Aldrich

MOPS, anhydrous, free-flowing, Redi-Dri^™, ≥99.5%

PHR1330

Supelco

Monobasic Potassium Phosphate, Pharmaceutical Secondary Standard; Certified Reference Material

1551139

USP

Monobasic potassium phosphate, United States Pharmacopeia (USP) Reference Standard

T6074

Sigma-Aldrich

Anti-α-Tubulin antibody, Mouse monoclonal, clone B-5-1-2, purified from hybridoma cell culture

229806

Sigma-Aldrich

Potassium phosphate monobasic, 99.99% trace metals basis

03609

Sigma-Aldrich

Ethylenediaminetetraacetic acid, BioUltra, ≥99.0% (KT)

03620

Sigma-Aldrich

Ethylenediaminetetraacetic acid, ≥98.0% (KT)

60218

Sigma-Aldrich

Potassium phosphate monobasic, BioUltra, for molecular biology, anhydrous, ≥99.5% (T)

03690

Sigma-Aldrich

Ethylenediaminetetraacetic acid disodium salt solution, BioUltra, for molecular biology, pH 8.0, ~0.5 M in H₂O

60229

Sigma-Aldrich

Potassium phosphate monobasic, tested according to Ph. Eur., anhydrous

E6758

Sigma-Aldrich

Ethylenediaminetetraacetic acid, anhydrous, crystalline, BioReagent, suitable for cell culture

E9884

Sigma-Aldrich

Ethylenediaminetetraacetic acid, ACS reagent, 99.4-100.6%, powder

P9791

Sigma-Aldrich

Potassium phosphate monobasic, for molecular biology, ≥98.0%

P5379

Sigma-Aldrich

Potassium phosphate monobasic, ReagentPlus^®

P5655

Sigma-Aldrich

Potassium phosphate monobasic, powder, suitable for cell culture, suitable for insect cell culture, suitable for plant cell culture, ≥99.0%

E8008

Sigma-Aldrich

Ethylenediaminetetraacetic acid solution, 0.02% in DPBS (0.5 mM), sterile-filtered, BioReagent, suitable for cell culture

Sigma-Aldrich

Ethylenediaminetetraacetic acid, purified grade, ≥98.5%, powder

EDS

Sigma-Aldrich

Ethylenediaminetetraacetic acid, anhydrous, BioUltra, ≥99% (titration)

431788

Sigma-Aldrich

Ethylenediaminetetraacetic acid, 99.995% trace metals basis