Briesacher S L, May T, Grigsby K N, Kerley M S, Anthony R V, Paterson J A
University of Missouri, Columbia 65211.
J Anim Sci. 1992 Jan;70(1):289-95. doi: 10.2527/1992.701289x.
We used DNA probes to study dietary effects on the prokaryotic population in the rumen. Procedures used to isolate and quantify prokaryotic 16S ribosomal RNA (rRNA) from the rumen using universal and species-specific DNA probes were evaluated. In this experiment, three ruminally fistulated steers were fed orchard-grass hay, and ruminal digesta were collected at 0, 3, and 9 h after offering hay (0800). Samples of ruminal digesta were taken from the interior portion of the digesta mat and from the fluid below the mat in the dorsal rumen. Freezing (-65 degrees C) and blending samples both increased (P less than .07) the yield of 16S rRNA from ruminal digesta. Extraction of prokaryotic rRNA was greater (P less than .04) when phenol buffered with sodium acetate was used than when it was buffered with hydroxymethyl-amino-methane. Prokaryotic 16S rRNA concentration of the fluid phase was similar (P greater than .10) at 0, 3, and 9 h after offering hay. Prokaryotic 16S rRNA concentration of the mat phase increased up to the 9 h after feeding. The proportion of Fibrobacter succinogenes remained constant in both digesta phases at all times measured. From these data we concluded that DNA probes can be used to monitor bacterial population shifts in the rumen.
我们使用DNA探针来研究日粮对瘤胃原核生物群体的影响。对使用通用和种特异性DNA探针从瘤胃中分离和定量原核生物16S核糖体RNA（rRNA）的程序进行了评估。在本试验中，给3头装有瘤胃瘘管的阉牛饲喂果园草干草，并在饲喂干草（08:00）后0、3和9小时采集瘤胃消化物。瘤胃消化物样本取自消化物垫的内部以及背侧瘤胃中垫子下方的液体。冷冻（-65℃）和混合样本均提高了（P＜0.07）瘤胃消化物中16S rRNA的产量。当使用用醋酸钠缓冲的苯酚时，原核生物rRNA的提取量比用羟甲基氨基甲烷缓冲时更高（P＜0.04）。饲喂干草后0、3和9小时，液相中原核生物16S rRNA浓度相似（P＞0.10）。垫子相中，原核生物16S rRNA浓度在采食后9小时内持续上升。在所有测定时间，两个消化物相中琥珀酸丝状杆菌的比例均保持恒定。根据这些数据，我们得出结论，DNA探针可用于监测瘤胃中细菌群体的变化。
