本文選題：高通量測(cè)序 + 水稻土��； 參考：《土壤學(xué)報(bào)》2017年01期

【摘要】：干濕交替是自然界普遍存在的現(xiàn)象,但長(zhǎng)期以來(lái)由于技術(shù)的限制,復(fù)雜土壤中微生物對(duì)水分變化的響應(yīng)規(guī)律仍不清楚。針對(duì)我國(guó)江蘇常熟湖泊底泥發(fā)育的典型水稻土,在室內(nèi)開展?jié)駶?rùn)-風(fēng)干以及風(fēng)干-濕潤(rùn)各三次循環(huán),每次循環(huán)中濕潤(rùn)、風(fēng)干狀態(tài)各維持7d,利用微生物核糖體rRNA的通用引物進(jìn)行PCR擴(kuò)增,通過(guò)高通量測(cè)序分析土壤古菌多樣性變化,同時(shí)結(jié)合實(shí)時(shí)熒光定量PCR技術(shù),在DNA和RNA水平研究古菌數(shù)量對(duì)干濕交替過(guò)程的響應(yīng)規(guī)律。結(jié)果表明:水稻土濕潤(rùn)-風(fēng)干過(guò)程中,在DNA水平土壤古菌數(shù)量降幅約為149倍~468倍,而在RNA水平降幅最高僅為2.06倍;水稻土風(fēng)干-濕潤(rùn)過(guò)程中,在DNA水平古菌數(shù)量增幅在147倍~360倍之間,而在RNA水平增幅最高僅為2.95倍。表明在干濕交替過(guò)程中,DNA水平的古菌16S rRNA基因數(shù)量變化遠(yuǎn)高于RNA水平�；诟咄繙y(cè)序多樣性的結(jié)果表明,在DNA和RNA水平,濕潤(rùn)土壤3次風(fēng)干、以及風(fēng)干土壤3次加水濕潤(rùn)7d恢復(fù)后,土壤古菌群落結(jié)構(gòu)均發(fā)生統(tǒng)計(jì)顯著性改變。在微生物門、綱、目、科和屬的不同分類水平下,水稻土古菌主要包括3、10、13、14、10種不同的類群,在RNA和DNA水平的結(jié)果基本一致。干濕交替導(dǎo)致部分古菌類群發(fā)生顯著變化,其中在微生物分類學(xué)目水平發(fā)生顯著變化的古菌最高達(dá)到6種,主要包括產(chǎn)甲烷古菌和氨氧化古菌,如Methanobacteriales、Methanosarcinales、Methanomicrobiales和Nitrososphaerales等。這些研究結(jié)果表明,反復(fù)的干濕交替并未顯著改變水稻土中古菌的主要類群組成,古菌類群的絕對(duì)數(shù)量和相對(duì)豐度發(fā)生了一定程度的變化,但這些變化與微生物生理作用的聯(lián)系仍需進(jìn)一步研究;風(fēng)干土壤中古菌RNA序列極可能來(lái)自于完整的古菌細(xì)胞,暗示了這些古菌細(xì)胞能夠較好地適應(yīng)水稻土中水分的劇烈變化,風(fēng)干狀態(tài)的土壤在一定程度也可用于土壤古菌群落組成研究。
[Abstract]:Dry-wet alternation is a common phenomenon in nature, but for a long time due to the limitation of technology, the response of microorganisms to water change in complex soil is still unclear. In view of typical paddy soils developed in the sediments of lakes in Changshu, Jiangsu Province, three cycles of wet-air drying and air-dry-wetting were carried out indoors, and each cycle was wetted. The diversity of soil palaeomycetes was analyzed by using universal primers of microbial ribosomal rRNA for 7 days, and the diversity of soil palaeomycetes was analyzed by high-throughput sequencing. Meanwhile, real-time fluorescence quantitative PCR technique was used to analyze the diversity of soil palaeomycetes. The response of ancient bacteria quantity to the alternating process of dry and wet was studied at DNA and RNA levels. The results showed that in the process of wetting and air drying of paddy soil, the decrease of soil paleomycetes at DNA level was about 149 times and 468 times, while that at RNA level was only 2.06 times, and in the process of air-drying and wetting of paddy soil, the decrease of palaeomycetes in paddy soil was about 149-fold and 2.06 times, respectively. The number of palaeomycetes increased by 147-fold in DNA level, but only 2.95 times in RNA level. The results showed that the change of 16s rRNA gene number of Archaea was much higher than that of RNA during dry-wet alternation. Based on the diversity of high throughput sequencing, the results showed that the community structure of soil paleomycetes changed statistically significantly at DNA and RNA levels, 3 times of dry soil and 3 times of dry soil after 7 days of water wetting. Under the different taxonomic levels of phylum, class, order, family and genus, there are mainly 10 different taxa of Paddy soil ancient bacteria, which are the same at RNA and DNA levels. The alternation of dry and wet resulted in significant changes of some paleomycetes, of which the highest was 6 species at the level of microbial taxonomic order, mainly including methanobacterium palaeomycetes and ammonia-oxidizing ancient bacteria, such as Methanobacteriae alesus Methanosarcinales method anomicrobiales and Nitrososphaerales. These results showed that repeated dry-wet alternation did not significantly change the composition of the main taxa in paddy soil, and the absolute number and relative abundance of paleomycetes changed to a certain extent. However, the relationship between these changes and the physiological effects of microbes still needs further study. The RNA sequence of palaeomycetes in air-dried soil is likely to come from intact paleoblast cells, suggesting that these paleomycetes cells can adapt to the severe changes of water in paddy soil. The dry soil can also be used to study the composition of soil paleomycetes community to some extent.
【作者單位】： 中國(guó)科學(xué)院南京土壤研究所;中國(guó)科學(xué)院大學(xué);
【基金】：國(guó)家重點(diǎn)基礎(chǔ)研究發(fā)展計(jì)劃(973計(jì)劃)項(xiàng)目(2015CB150501)資助~~
【分類號(hào)】：S154.3
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本文編號(hào)：1962207