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产品介绍

微生物是一群以分解代谢为主的生物类群,其生物活性十分丰富。16S rDNA基因测序已经成为当前解析原核微生物群落组成及其分布的重要手段。既往研究中,受限于NGS的读长,物种鉴定存在很多不确定性。伴随着三代测序技术的发展,长度长测序技术越来越成为组学研究的主流。Pacbio平台3-5kb的平均读长可以轻而易举地覆盖16s rDNA9个高变区域,获得的序列更长,信息量也更多!16s rDNA全长测序因此成为多样性研究的新热点技术。

基于三代PacBio高通量技术对微生物16S rDNA(真菌ITS、真核生物18S rRNA)全长序列进行测序,物种能够鉴定到“种”水平,同时对样品中的优势物种、稀有物种以及一些未知的物种进行检测,获得样品中的微生物群落组成以及它们之间的相对丰度。探讨微生物多样性对于研究微生物与环境的关系、环境治理和微生物资源的利用有着重要的理论和现实意义。此外,凌恩生物还推出个性化分析服务,如:筛选微生态系统中的关键菌种、微生物群落变化的动态监测、微生物群落共生网络关系等多项个性化分析服务,为您的科研成果锦上添花。


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技术优势

1基于三代测序,在同一分类水平下,也可以实现更多的物种分类,更是在种水平物种鉴定中实现了大幅提升,这一点就很大程度满足了微生物学研究者的迫切需求。

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16s V4区与全长分类鉴定结果比较

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16s不同V区与全长未分类序列占比比较

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16s不同V区与全长未分类序列进化树比较


2基于三代技术开展16s全长扩增子研究,有更多的序列得到了明确的物种划分,尤其是针对样本中含量低微的物种,分类鉴定的效果更好。

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样本中微量物种的分类识别情况和偏好性16s二代和三代比较


3由于二代测序建库过程中有PCR扩增的步骤,难以避免由于扩增偏好性造成的物种丰度检测的偏差,可能造成某些特定微生物类群含量的检测偏差,这就难以实现对样本中微生物群落分布的真实还原。而三代测序由于不需要PCR的过程,就很大程度上降低了结果的偏差

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16s V4区与全长对物种偏好性比较


4通过聚类OTU,并与模拟物种库进行比较,V1-V9区研究实现了对样本的更真实还原,物种的识别和定量是最接近模拟物种库原貌的,这一点说明了基于三代平台的16s全长测序在保真效果上的巨大优势

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16s不同V区与全长与模拟物种库进行比较


5利用三代16s全长扩增子测序技术可以清晰的获得菌株在16s全长范围内的单核苷酸变异信息。利用这样的信息,可以更好地对菌株实现区分物种可实现“种”水平分类,结果更加准确

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技术流程

PacBio测序(扩增子全长测序):

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内容项目引物名称序列扩增产物长度
全长16S全长27F5′-AGRGTTTGATYNTGGCTCAG-3′1.5K
1492R5′-TASGGHTACCTTGTTASGACTT-3′
18S全长EukA5′-AACCTGGTTGATCCTGCCAGT-3′1.8K
EukB5′-GATCCTTCTGCAGGTTCACCTAC-3
ITS全长ITS15′- CTTGGTCATTTAGAGGAAGTAA-3′0.6-0.7K
ITS45′- TCCTCCGCTTATTGATATGC-3′



分析流程

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送样要求


样本类型1:环境样本(请使用干冰或者冰袋运送)

1)土壤≥5g

2)粪便≥2g

样本类型2DNA样品(请使用干冰或者冰袋运送)

1DNA总量1ug

2)浓度20ng/ul



分析示例


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部分凌恩客户文章


发表时间

文章标题

杂志名

影响因子

合作单位

2023

Unearthing the mechanisms underlying calcium carbonate therapies for eliminating pathogens during composting

Chemical Engineering Journal

15.1

中国科学院

2022

ETS-1 facilitates Th1 cell-mediated mucosal inflammation in inflammatory bowel diseases through upregulating CIRBP

Journal of Autoimmunity

14.511

同济大学

2023

Magnolol-driven microbiota modulation elicits changes in tryptophan metabolism resulting in reduced skatole formation in pigs

JoualofHazardous Materials

13.6

广东省农业科学院

2022

Bacterial dynamics of drinking water pipeline: A new perspective on dual effects of stagnation and algal organic matter

Water Research

13.4

西安建筑技术大学

2022

Bacterial community structure and metabolic activity of drinking water pipelines in buildings: A new perspective on dual effects of hydrodynamic stagnation and algal organic matter invasion

Water Research

13.4

西安建筑科技大学

2024

Vermicomposting preferably alters fungal communities in wasted activated sludge and promotes the production of plant growth-promoting biostimulants in the vermicompost

Chemical Engineering Journal

13.3

中国地质大学

2024

Self-circulating pulse alternating ventilation composting technology: Biotic and abiotic effects of mature compost on humification during food waste composting

Chemical Engineering Journal

13.3

浙江大学

2023

Interaction of silver nanoparticles with marine/lake snow in early formation stage

Water Research

12.8

华中科技大学

2024

Polystyrene nanoplastics alter intestinal toxicity of 2,4-DTBP in a sex-dependent manner in zebrafish (Danio rerio)

Journal of Hazardous Materials

12.2

内蒙古大学

2025

Algal organic matter triggers re-assembly of bacterial community in plumbing system

Journal of Hazardous Materials

12.2

西安建筑科技大学

2024

Priority establishment of soil bacteria in rhizosphere limited the spread of tetracycline resistance genes from pig manure to soil-plant systems based on synthetic communities approach

Environment International

11.8

华南农业大学

2024

Human microbiota from drug-naive patients with obsessive-compulsive disorder drives behavioral symptoms and neuroinflammation via succinic acid in mice

Molecular Psychiatry

11

上海交通大学

2025

Potent and Selective Inhibition of Sulfate-Reducing Bacteria by Neutral Red

Environmental Science & Technology

10.9

哈尔滨工业大学

2022

Small Intestine-specific Knockout of CIDEC Improves Obesity and Hepatic Steatosis by Inhibiting Synthesis of Phosphatidic Acid

Int. J. Biol. Sci

10.75

广西大学

2023

Cadmium-resistant phosphate-solubilizing bacteria immobilized on phosphoric acid-ball milling modified biochar enhances soil cadmium passivation and phosphorus bioavailability

Science of the Total Environment

9.8

四川农业大学

2023

Polymeric immunoglobulin receptor deficiency exacerbates autoimmune hepatitis by inducing intestinal dysbiosis and barrier dysfunction

Cell Death and Disease

9.685

温州医科大学

2023

Degradation potential and pathways of methylcyclohexane by bacteria derived from Antarctic surface water

Chemosphere

8.943

海洋生物资源与环境研究中心

2024

Degradation and potential metabolism pathway of polystyrene by bacteria from landfill site

Environmental Pollution

8.9

北京科技大学

2023

Synthesis of an autochthonous microbial community by analyzing the core microorganisms responsible for the critical flavor of bran vinegar

Food Research International

8.1

四川轻化工大学

2023

Monitoring and identification of spoilage-related microorganisms in braised chicken with modified atmosphere packaging during refrigerated storage

Food Science and Human Wellness

8.022

南京农业大学

2024

Vitamin K2 alleviates dextran sulfate sodium-induced colitis via inflammatory responses, gut barrier integrity, and the gut microbiota in mice

International Journal of Biological Macromolecule

7.7

安徽省农业科学院

2025

Protein hydrolysate from Atlantic salmon (Salmo salar) improves aging-associated neuroinflammation and cognitive decline in rats by reshaping the gut microbiota and Th17/Treg balance

International Journal of Biological Macromolecules

7.7

山西医科大学

2024

Gut microbiota-based discovery of Houttuyniae Herba as a novel prebiotic of Bacteroides thetaiotaomicron with anti-colitis activity

Biomed Pharmacother

7.5

天津中医药大学

2024

Structural changes and assembly mechanisms of microbial communities during rapid sedimentation of Yellow River sediments

Environmental Technology & Innovation

7.1

济南大学

2024

Untargeted metabolomics and PacBio analysis on bioactive components and microbial community in co-fermentation of black soldier fly larva

Food Research International

7

热带农业科学院

2023

Anti-virulence activity of novel (1-heteroaryloxy-2- hydroxypropyl)- phenylpiperazine derivatives against both wild-type and clinical drug-resistant Candida albicansstrains

Microbial Biotechnology

6.575

广州医科大学

2022

Linarin ameliorates dextran sulfate sodium-induced colitis in C57BL/6J mice via the improvement of intestinal barrier, suppression of inflammatory responses and modulation of gut microbiota

Food Funct.

6.317

西北农林科技大学

2022

Heat Stress Weakens the Skin Barrier Function in Sturgeon by Decreasing Mucus Secretion and Disrupting the Mucosal Microbiota.

Frontiers in Microbiology

6.064

四川农业大学

2022

Characterization of the spoilage heterogeneity of Aeromonas isolated from chilled chicken meat: in vitro and in situ

Lebensmittel-Wissenschaft & Technologie

6.056

南京农业大学

2022

The Gut Microbial Co-Abundance Gene Groups (CAGs) Differentially Respond to the Flavor (Yao-Wei) of Chinese Meteria Medica

AM J CHINESE MED

6.005

天津中医药大学