Effects of fescue cultivar and heat shock protein haplotype on growth ...

arkansasagnews.uark.edu

Effects of fescue cultivar and heat shock protein haplotype on growth ...

AAES Research Series 5971,200 × g for 20 min (Marathon 22KBR, Fisher Scientific, Hermle-Labortechnik, Germany) to isolate the buffy coat. Genomic DNA wasobtained from the buffy coat (QIAGEN Inc., Valencia, Calif.) ong>andong>diluted to 20 ng/µL.Heifers were ong>haplotypeong>d based on Hsp70 promoter sequenceas described by Rosenkrans et al. (2010); No = no SNP; Deletion =cytosine deletion at nucleotide base 895; Yes = a SNP other than adeletion. Heifers also were ong>haplotypeong>d for Hsp70 coding sequences;Haplotype 1 = same sequence as the National Center for BiotechnologyInformation (NCBI); Haplotype 2 = cytosine to guanine base changeat base 2087; ong>andong> Haplotype 3 = guanine to cytosine base change at base2033. Polymerase chain reaction (PCR) primers for Hsp70 promotersequencing [forward (5’-GCCAGGAAACCAGAGACAGA-3’) ong>andong>reverse (5’-CCTACGCAGGAGTAGGTGGT-3’)] were utilized toamplify a 539-base sequence that spanned a conserved region withinthe Hsp70 promoter region. Coding sequence primers for Hsp70[forward (5’-GAAGAGCGCCGTGGAGGATG -3’) ong>andong> reverse(5’-CTTGGAAGTAAACAGAAACGGG -3’)] were utilized toamplify a 570-base sequence for PCR. A Peltier thermal cycler (MJResearch, Waltham, Mass.) was used for PCR. Each PCR began withan initial 2-min ong>heatong>ing at 94 °C, followed by 35 cycles at 94 °C for 30s, 1 min at 55 °C, ong>andong> 1 min at 68 °C. A final elongation step consistedong>ofong> 10 min at 68 °C. Approximately 5 µL ong>ofong> 20 ng/µL genomic DNA,1 µL ong>ofong> 10 µM forward primer, 1 µL ong>ofong> 10 µM ong>ofong> reverse primer ong>andong>43 µL ong>ofong> Platinum PCR Supermix (Invitrogen, Carlsbad, Calif.) fora total volume ong>ofong> 50 µL in each PCR. Amplicons were visualized viaelectrophoresis in 1.2% agarose gels stained with ethidium bromidein 1.0X Tris/Boric Acid/EDTA.Amplification products were purified using QIAquick PCRpurification kit (QIAGEN Inc., Valencia, Calif.). Purified PCRproducts were sequenced at the University ong>ofong> Arkansas DNA CoreLab using ABI Prism 3100 Genetic Analyzer (Applied Biosystems,Foster City, Calif.). Sequences were compared using the web-basedsong>ofong>tware package ClustalW (http://www.ebi.ac.uk/Tools/clustalw2/index.html; European Bioinformatics Institute, Cambridge, UK).Homozygous ong>andong> heterozygous alleles were identified by assessingsequence chromatograms using BioEdit.Statistical Analyses. The experimental design was rong>andong>omizedcomplete block ong>andong> pasture was the experimental unit in all statisticalanalyses. Haplotypes, growth measurements ong>andong> AFC were analyzedby ANOVA utilizing the MIXED procedure (SAS Inst., Cary, N.C.).Pregnancy rate was analyzed by chi-squared analysis ong>ofong> SAS.Results ong>andong> DiscussionHeifers grazing E+ tall ong>fescueong> had reduced overall growthcompared to heifers grazing Novel ong>fescueong> which reiterates previousresearch (Paterson et al., 1995). Overall ADG tended (P = 0.08) tobe greater for heifers grazing novel (1.32 ± 0.22 lbs) than E+ heifers(0.88 ± 0.22 lbs). Heifers grazing E+ tall ong>fescueong> had smaller pelvicheight, pelvic width ong>andong> pelvic area. Hip width increased (day effect;(P < 0.001) over time. Novel heifers (14.4 ± 0.1 in) had larger (P 0.12) between ong>haplotypeong>s ong>andong>ong>fescueong> or ong>haplotypeong>s ong>andong> day. Heifers grazing E+ tall ong>fescueong> had higherAFC compared to Novel heifers; however, Novel heifer pregnancyrates were greater than heifers grazing E+. The day ong>ofong> scan affected (P< 0.007) AFC (9.0 antral follicles, 13.6 antral follicles, ong>andong> 10.2 antralfollicles; d 75, 85, ong>andong> 183, respectively) (Table 2). Fescue ong>cultivarong>affected (P < 0.002) AFC (8.4 antral follicles vs. 13.5 antral follicles;Novel vs.E+) ong>andong> pregnancy rates (81.0% vs. 36.0%; Novel vs.E+).These data are inconsistent with previous studies that indicate AFCis associated with increased fertility in cattle (Irelong>andong> et al., 2008);however, there may be other factors involved. Haplotypes ong>ofong> Hsp70promoter (P < 0.08) ong>andong> coding sequence (P < 0.003) affected AFC.ImplicationsHeifers grazing toxic endophyte tall ong>fescueong> had a reduction inoverall growth compared to heifers grazing novel endophyte infectedong>fescueong>; thus, livestock producers should utilize novel endophyteinfected ong>fescueong> for development ong>ofong> replacement beef heifers. Heifersgrazing toxic endophyte infected ong>fescueong> had higher antral folliclecounts compared to novel endophyte heifers; however, pregnancy ratein heifers grazing novel endophyte infected ong>fescueong> was significantlyhigher than in heifers grazing toxic endophyte infected ong>fescueong>suggesting other factors may have impacted antral follicle count.Additionally, ong>heatong> ong>shockong> ong>proteinong> 70 promoter ong>andong> coding sequenceong>haplotypeong>s may be useful indicators ong>ofong> fertility in replacement heifers.AcknowledgementsThis study was supported in part by USDA, ARS cooperativeagreement #58-6227-8-042. The authors would like to thank BrentWoolley ong>andong> Sam Tabler, USDA-ARS for technical support ong>andong> dailyanimal management.Literature CitedBurke, J. M., R. W. Rorie, E. L. Piper, ong>andong> W. G. Jackson. 2001.Reproductive responses to grazing endophyte-infected tall ong>fescueong>by post-partum beef cows. Therio.56:357-369.Burke, J. M., D. E. Spiers, F. N. Kojima, G. A. Perry, B. E. Salfen, S. L.Wood, D. J. Patterson, M. F. Smith, M. C. Lucy, W. G. Jackson,ong>andong> E. L. Piper. 2004. Interaction ong>ofong> endophyte-infected ong>fescueong>ong>andong> ong>heatong> stress on ovarian function in the beef heifer. Biol.Reprod.65:260-268.Irelong>andong>, J. L. H., D. Scheetz, F. Jimenez-Krassel, A. P. N. Themmen, F.Ward, P. Longeran, G. W. Smith, G. I. Perez, A. C. O. Evans, ong>andong> J.J. Irelong>andong>. 2008. Antral follicle count reliability predicts number ong>ofong>morphologically healthy oocytes ong>andong> follicles in ovaries ong>ofong> youngadult cattle. Biol. Reprod.79:1219-1225.Paterson, J., C. Forcherio, B. Larson, M. Samford, ong>andong> M. Kerley. 1995.The effects ong>ofong> ong>fescueong> toxicosis on beef cattle productivity. J. Anim.Sci.73:889-898.Rosenkrans, C. F., Jr., A. Banks, S. Reiter, ong>andong> M. L. Looper. 2010.Calving traits ong>ofong> crossbred Brahman cows are associated withHeat Shock Protein 70 genetic polymorphisms. Anim. Reprod.Sci.119:178-182.58


Arkansas Animal Science Department Report 2011Table 1. Overall growth ong>andong> development measurements ong>ofong> beef heifersgrazing either toxic endophyte-infected (E+) or non-toxic (Novel)endophyte-infected tall ong>fescueong>.Parameter E+ Novel P ValueAverage Daily Gain, lbs 0.88 ± 0.22 1.32 ± 0.22 0.08Hip Width, in 14.1 ± 0.1 14.4 ± 0.1 < 0.05Pelvic Height, in 5.3 ± 0.1 5.5 ± 0.1 < 0.06Pelvic Area, in 2 58.6 ± 1.2 61.6 ± 1.2 < 0.06Exit Velocity, s/m 0.6 ± 0.1 0.8 ± 0.1 < 0.08Table 2. Heifer prebreeding antral follicle count (AFC) ong>andong> subsequentpregnancy rate ong>ofong> beef heifers grazing either toxic endophyte-infected (E+)or non-toxic (Novel) endophyte-infected tall ong>fescueong>.Parameter E+ Novel P ValueAntral Follicle Count 13.7 10 < 0.002Pregnancy Rate, % 36.0 81.0 < 0.0559

More magazines by this user
Similar magazines