Desulfotomaculum genus- and subgenus-specific 16S rRNA hybridization probes for environmental studies.
Desulfotomaculum genus- and subgenus-specific 16S rRNA hybridization probes for environmental studies.
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用于环境研究的 Desulfotomaculum 属和亚属特异性 16S rRNA 杂交探针。
DOI：
10.1046/j.1462-2920.2000.00085.x
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发表时间：
2000
影响因子：
5.1
通讯作者：
Raskin,L
中科院分区：
文献类型：
--
作者：
Hristova,KR;Mau,M;Zheng,D;Aminov,RI;Mackie,RI;Gaskins,HR;Raskin,L
根据16 SrRNA序列的比较分析和最近建立的脱硫肠状菌属的遗传学基础，开发了一套遗传学巢式杂交探针。一种属特异性探针靶向所有已知的脱硫肠状菌属（除乙酰氧化脱硫肠状菌外），五种特异性探针靶向脱硫肠状菌属内的亚群。通过实验确定每个探针的解离温度。通过与从各种靶和非靶微生物中分离的纯培养rRNA杂交，并通过使用从四种不同环境中获得的样本评价探针“嵌套”，验证了探针特异性。还优化了荧光原位杂交的固定和杂交条件。用该探针进行膜杂交定量分析，结果表明，在高温厌氧消化池、土壤、人粪便和猪结肠中，脱硫肠杆菌rRNA分别占总rRNA的0.3-2.1%、2.6-6.6%、1.5-3.3%和2.5-6.2%。
Based on comparative analysis of 16S rRNA sequences and the recently established phylogeny of the genusDesulfotomaculum, a set of phylogenetically nested hybridization probes was developed and characterized. A genus‐specific probe targets all knownDesulfotomaculumspecies (with the exception ofDesulfotomaculum acetoxidans), and five specific probes target subclusters within theDesulfotomaculumgenus. The dissociation temperature of each probe was determined experimentally. Probe specificities were verified through hybridizations with pure culture rRNA isolated from a wide variety of target and non‐target organisms and through an evaluation of probe ‘nesting’ using samples obtained from four different environments. Fixation and hybridization conditions for fluorescencein situhybridizations were also optimized. The probes were used in quantitative membrane hybridizations to determine the abundance ofDesulfotomaculumspecies in thermophilic anaerobic digesters, in soil, in human faeces and in pig colon samples.DesulfotomaculumrRNA accounted for 0.3–2.1% of the total rRNA in the digesters, 2.6–6.6% in soil, 1.5–3.3% in human faeces and 2.5–6.2% in pig colon samples.