China Metagenomic Sequencing (NGS) umzi-mveliso kunye nabavelisi |BMK

Ulandelelwano lweMetagenomic (NGS)

Metagenomic Sequencing (NGS) Featured Image

I-Metagenome ibhekiselele kwingqokelela yemathiriyeli yemfuzo iyonke yoluntu oluxubeneyo lwezinto eziphilayo, ezifana ne-metagenome yokusingqongileyo, i-metagenome yabantu, njl njl.Ukulandelelana kwe-Metagenomic sisixhobo se-molecular esetyenziselwa ukuhlalutya izinto ezixubeneyo ze-genomic ezithathwe kwiisampuli zendalo, ezinikezela ngolwazi olucacileyo kwiintlobo zeentlobo zeentlobo kunye nobuninzi, ubume benani labantu, ubudlelwane be-phylogenetic, izakhi zofuzo ezisebenzayo kunye nenethiwekhi yokulungelelanisa kunye nezinto ezisingqongileyo.

Iqonga:Illumina NovaSeq6000

Qhagamshelana nathi

Iinkcukacha zeNkonzo

Iziphumo zeDemo

Into yokuphonononga

Iinkonzo eziluncedo

ØUkwahlukaniswa kunye nokulinywa simahla kwiprofayili yoluntu lwemicrobial

ØIsisombululo esiphezulu ekufumaneni iintlobo eziphantsi kobuninzi kwiisampuli zokusingqongileyo

ØIngcamango ye "meta-" idibanisa zonke iimpawu zebhayoloji kwinqanaba lokusebenza, inqanaba leentlobo kunye nenqanaba lemfuza, ebonisa umbono oguqukayo osondeleyo kwinyani.

ØI-BMK iqokelela amava amakhulu kwiintlobo zeesampulu ezahlukeneyo ezineisampulu ezingaphezulu kwe-10,000 ezicutshungulweyo.

IiNgcaciso zeNkonzo

UkulandelelanisaIqonga	Ithala leencwadi	Imveliso yedatha ecetyiswayo	Ixesha eliqikelelweyo lokujika
Illumina NovaSeq 6000	PE250	50K/100K/300K iithegi	Iintsuku ezingama-30

Uhlalutyo lwe-Bioinformatics

üUlawulo lomgangatho wedatha ekrwada

üIndibano yeMetagenome

üIseti yemfuza engafunekiyo kunye nenkcazo

üUhlalutyo lweentlobo ezahlukeneyo zeentlobo

üUhlalutyo lokwahluka komsebenzi wemfuza

üUhlalutyo lwamaqela

üUhlalutyo lombutho ngokuchasene nezinto zovavanyo

IiMfuno zeSampulu kunye nokuhanjiswa

IiMfuno zeSampulu:

KubaIzicatshulwa zeDNA:

Uhlobo lweSampuli	Isixa	Ukugxila	Ubunyulu
Izicatshulwa zeDNA	＞ 30 ng	＞ 1 ng/μl	OD260/280= 1.6-2.5

Ngeesampulu zokusingqongileyo:

Uhlobo lwesampuli	Inkqubo yesampulu ecetyiswayo
Umhlaba	Isixa sesampulu: malunga.5 g;Into eseleyo ebunayo kufuneka isuswe kumphezulu;Gaya iinqununu ezinkulu kwaye udlule kwi-2 mm filter;Iisampulu ze-Aliquot kwityhubhu ye-EP okanye i-cyrotube yogcino.
Ilindle	Isixa sesampulu: malunga.5 g;Qokelela kwaye iisampulu ze-aliquot kwi-EP-tube engcolileyo okanye i-cryotube yokubhukisha.
Iziqulatho zamathumbu	Iisampuli kufuneka ziqwalaselwe phantsi kwemeko ye-aseptic.Hlamba izicubu eziqokelelweyo nge-PBS;I-Centrifuge i-PBS kwaye uqokelele i-precipitant kwi-EP-tubes.
Udaka	Isixa sesampulu: malunga.5 g;Qokelela kwaye isampuli ye-aliquot yodaka kwi-EP-tube engcolileyo okanye i-cryotube yokugcinelwa
Amanzi	Kwisampulu enesixa esilinganiselweyo se-microbial, njengamanzi epompo, amanzi omthombo, njl., Qokelela ubuncinci be-1 L yamanzi kwaye udlule kwi-0.22 μm yokucoca ukutyebisa i-microbial kwi-membrane.Gcina inwebu kwityhubhu enyumba.
Ulusu	Khulula ngononophelo umphezulu wolusu ngeswab yomqhaphu engcolileyo okanye ngencakuba yotyando kwaye uyibeke kwityhubhu engenanto.

Unikezelo lweSampuli eCetyisiweyo

Umkhenkce iisampulu kulwelo nitrogen iiyure 3-4 kwaye ugcine kulwelo nitrogen okanye -80 degree ukuya ugcino ixesha elide.Isampulu yokuthumela ngenqanawe eyomileyo iyafuneka.

Ukuhamba komsebenzi weNkonzo

Ukuhanjiswa kwesampuli

Ukwakhiwa kwethala leencwadi

Ukulandelelanisa

Uhlalutyo lwedatha

Iinkonzo emva kokuthengisa

Ngaphambili: Ulandelelwano lweMetagenomic-Nanopore

Okulandelayo: I-Chromatin Immunoprecipitation Sequencing (ChIP-seq)

1.I-Histogram: Ukuhanjiswa kweentlobo

2.Ufuzo olusebenzayo oluchazwe kwiindlela ze-metabolic ze-KEGG

3.Imephu yobushushu: Imisebenzi eyohlukileyo esekelwe kubuninzi bemfuzaI-4.I-Circos ye-CARD i-antibiotic resistance genenes

Ityala le-BMK

Ukuxhaphaka kofuzo lokuxhathisa amayeza okubulala iintsholongwane kunye neentsholongwane zebhaktiriya ecaleni kwengcambu yommango

Ipapashiwe:Ijenali yeZixhobo ezinobungozi, ngo-2021

Iqhinga lokulandelelanisa:

Izinto: Izicatshulwa ze-DNA zamaqhekeza amane eengcambu zemangrove ezinxulumene neesampuli: umhlaba ongatyalwanga, irhizosphere, i-episphere kunye ne-endosphere compartments
Iqonga: Illumina HiSeq 2500
Ujoliso: iMetagenome
16S rRNA gene V3-V4 ummandla

Iziphumo eziphambili

Ulandelelwano lwe-Metagenomic kunye ne-metabarcoding profiling kwi-root-root continuum ye-mangrove saplings yacutshungulwa ukuze kufundwe ukusasazwa kofuzo lwe-antibiotic resistance (ARGs) ukusuka emhlabeni ukuya kwizityalo.Idatha ye-Metagenomic ibonise ukuba i-91.4% yejene yokumelana namayeza okubulala iintsholongwane aye achongwa ngokuqhelekileyo kuwo onke amacandelo omhlaba akhankanywe ngasentla, abonisa ifashoni eqhubekayo.Ulandelelwano lwe-amplicon ye-16S rRNA yenze i-29,285 yokulandelelana, emele iintlobo ze-346.Ukudibanisa neentlobo zeprofayili ngokulandelelana kwe-amplicon, oku kusasazwa kwafunyaniswa kuzimeleyo kwi-microbiota ehambelana neengcambu, nangona kunjalo, inokuququzelelwa yi-mobile ye-genetic elements.Olu phononongo luchonge ukuhamba kwee-ARG kunye neentsholongwane ezibangela izifo ezisuka emhlabeni zisiya kwizityalo ngokudityaniswa kweengcambu zomhlaba.

Isalathiso

Wang, C. , Hu, R. , Strong, PJ , Zhuang, W. , & Shu, L. .(2020).Ukuxhaphaka kwejini yokuxhathisa amayeza okubulala iintsholongwane kunye neentsholongwane ezibangela iintsholongwane ecaleni kwengcambu yommango.Ijenali yeZixhobo ezinobungozi, 408, 124985.