During January May 2009, severe polyarthritis was observed in ≈100% of ≈350 female calves at the central calf rearing unit of a farm in Helongjiang Province, People's Republic of China.
The DNA was then digested to nucleosides with 4 units of benzonase nuclease, 0.1 units phosphodiesterase 1, 4 units DNase, and 20 units calf intestine alkaline phosphatase in 10 mM Tris buffer (pH 7.9) and 1 mM magnesium chloride.
All cells were grown in 25 cm tissue culture flasks in RPMI-1640 with 25 mM HEPES supplemented with 2 mM l-glutamine, 15% fetal calf serum, 100 units penicillin, and 100 units streptomycin.
Calf ADG per unit of total feed energy intake was greater (P ≥ 0.05) for nursing pairs at ARDC but not different between EW and CW at PHREC, indicating early weaning may have minimal effect on reducing feed energy requirements.
The heel rise test is a common test of function of the calf muscle-tendon unit and is used as a test to assess calf muscle endurance and strength [ 5, 7, 8].
Digested DNA was dephosphorylated using 0.01 unit calf intestinal alkaline phosphatase per picomole ends of DNA at 50°C for 1 h, purified using an equal volume of 25:24:1 phenol chloroform:isoamyl alcohol, precipitated using 2.5 volumes of cold 100% ethanol and 3 M sodium acetate (NaOAc), and then resuspended in TE buffer (10 mM Tris [pH 8.0], 1 mM EDTA).
Briefly, DNA samples (4 μg) were digested with micrococcal nuclease (120 mUnits) and calf spleen phosphodiesterase (40 mUnits), enriched and labeled as reported elsewhere [Philips and Arlt, 2007].
DNA (4 μg) was digested with micrococcal nuclease (120 mUnits) and calf spleen phosphodiesterase (40 mUnits), enriched with Nuclease P1, and labeled with [γ-P]ATP as previously described [Phillips and Arlt, 2007].
HEK-293 cells (ATCC) were grown in MEM with 10% fetal calf serum, 100 units of penicillin/streptomycin, 2 mM L-glutamine, and 1 mM pyruvate.
All media were supplemented with 10% (v/v) fetal calf serum, 100 units penicillin and 100 µg/ml streptomycin, and grown at 37°C and 5% CO2.
Huh7 and 293T cells (American Type Culture Collection, Rockville, MD) were maintained in Dulbecco's Modified Eagle's Medium containing 10% heat-inactivated fetal calf serum, 100 Units of penicillin G sodium per ml, and 100 µg of streptomycin sulfate per ml (DMEM-10).
