Zogulitsa

Metagenomic Sequencing -NGS

Metagenome imatanthawuza kusonkhanitsa kwa chibadwa chamagulu osakanikirana a zamoyo monga chilengedwe, metagenome yaumunthu, ndi zina zotero. Lili ndi majeremusi a tizilombo tomwe timalima komanso tosalimidwa.Metagenomic sequencing ndi chida cha mamolekyu chomwe chimagwiritsidwa ntchito posanthula zinthu zosakanizika za ma genomic zotengedwa ku zitsanzo zachilengedwe, zomwe zimapereka chidziwitso chambiri pamitundu yosiyanasiyana komanso kuchuluka kwake, kuchuluka kwa anthu, ubale wa phylogenetic, majini ogwira ntchito komanso maukonde olumikizana ndi chilengedwe.

nsanja:Illumina NovaSeq Platform

Lumikizanani nafe

Tsatanetsatane wa Utumiki

Zotsatira za Demo

Nkhani Yophunzira

Ubwino wa Utumiki

● Kudzipatula komanso kulimidwa mosamalitsa kuti tipeze mbiri ya anthu ammudzi

● Kusamalitsa kwakukulu pozindikira mitundu yotsika kwambiri mu zitsanzo zachilengedwe

● Lingaliro la "meta-" limagwirizanitsa zinthu zonse zamoyo pamlingo wogwirira ntchito, mlingo wa mitundu ndi jini, zomwe zimasonyeza maonekedwe amphamvu omwe ali pafupi ndi zenizeni.

● BMK imakhala ndi zochitika zambiri zamitundu yosiyanasiyana ndi zitsanzo zoposa 10,000 zokonzedwa.

Mafotokozedwe a Utumiki

nsanja	Kutsata	Deta yovomerezeka	Nthawi yosinthira
Illumina NovaSeq Platform	Mtengo wa PE150	6G/10G/20G	45 Masiku ogwira ntchito

Bioinformatics kusanthula

● Kuwongolera khalidwe la data yaiwisi

● Msonkhano wa Metagenome

● Ma jini osagwiritsidwa ntchito mowonjezera ndi mawu ake

● Kusanthula zamitundu yosiyanasiyana

● Kusanthula kwa mitundu yosiyanasiyana ya ma genetic

● Kusanthula magulu

● Kusanthula kwamagulu motsutsana ndi zinthu zoyesera

Zitsanzo Zofunika ndi Kutumiza

Zitsanzo Zofunika:

ZaZithunzi za DNA:

Mtundu Wachitsanzo	Ndalama	Kukhazikika	Chiyero
Zithunzi za DNA	＞ 30 ng	＞ 1 ng/μl	OD260/280= 1.6-2.5

Za zitsanzo zachilengedwe:

Mtundu wachitsanzo	Njira yoyeserera yolangizidwa
Nthaka	Kuchuluka kwa zitsanzo: pafupifupi.5 g pa;Chotsalira chofota chiyenera kuchotsedwa pamwamba;Pewani zidutswa zazikulu ndikudutsa fyuluta ya 2 mm;Zitsanzo za Aliquot mu chubu chosabala cha EP kapena cyrotube kuti zisungidwe.
Ndowe	Kuchuluka kwa zitsanzo: pafupifupi.5 g pa;Sonkhanitsani ndi aliquot zitsanzo mu EP-chubu wosabala kapena cryotube kuti musungidwe.
Zomwe zili m'matumbo	Zitsanzo ziyenera kukonzedwa pansi pa chikhalidwe cha aseptic.Sambani minofu yosonkhanitsidwa ndi PBS;Centrifuge the PBS ndi kusonkhanitsa precipitant mu EP-machubu.
Sludge	Kuchuluka kwa zitsanzo: pafupifupi.5 g pa;Sungani ndi aliquot sludge zitsanzo mu wosabala EP-chubu kapena cryotube kusungitsa
Madzi	Kwa zitsanzo zokhala ndi tizilombo tating'onoting'ono, monga madzi apampopi, madzi a m'chitsime, ndi zina zotero, Sonkhanitsani madzi osachepera 1 L ndikudutsa 0.22 μm fyuluta kuti mulemeretse tizilombo toyambitsa matenda pa nembanemba.Sungani nembanemba mu chubu chosabala.
Khungu	Pala pamwamba pa khungu ndi thonje wosabala kapena tsamba la opaleshoni ndikuyika mu chubu wosabala.

Kutumiza Kwachitsanzo Kovomerezeka

Ikani zitsanzo mu nayitrogeni wamadzimadzi kwa maola 3-4 ndikusunga mu nayitrogeni wamadzimadzi kapena -80 digiri kuti musungidwe nthawi yayitali.Kutumiza kwachitsanzo kokhala ndi ayezi wowuma ndikofunikira.

Kuyenda kwa Ntchito Yantchito

Kupereka zitsanzo

Kumanga laibulale

Kutsata

Kusanthula deta

Pambuyo pogulitsa ntchito

Zam'mbuyo: Metagenomic Sequencing-Nanopore

Ena: Chromatin Immunoprecipitation Sequencing (ChIP-seq)

1.Histogram: Kugawa kwamitundu

2.Majini ogwira ntchito ofotokozera njira za metabolic za KEGG

Mapu a 3.Kutentha: Ntchito zosiyana zotengera kuchuluka kwa majini achibale4.Circos ya CARD antibiotic resistance genes

BMK Case

Kuchuluka kwa majini olimbana ndi maantibayotiki ndi tizilombo toyambitsa matenda m'mphepete mwa mizu ya mangrove

Lofalitsidwa:Journal of Hazardous Equipment, 2021

Njira yotsatirira:

Zida: DNA ya zidutswa zinayi za mizu ya mangrove yokhudzana ndi zitsanzo: nthaka yosabzalidwa, rhizosphere, episphere ndi endosphere compartments
Platform: Illumina HiSeq 2500
Zolinga: Metagenome
16S rRNA jini V3-V4 dera

Zotsatira zazikulu

Tsatanetsatane wa metagenomic ndi mbiri ya metabarcoding pamizu yopitilira muzu wa mitengo ya mangrove adakonzedwa kuti aphunzire kufalitsa ma antibiotic resistance genes (ARGs) kuchokera kunthaka kupita ku zomera.Deta ya Metagenomic idavumbulutsa kuti 91.4% ya majini olimbana ndi maantibayotiki amazindikirika m'zigawo zonse zinayi zadothi zomwe zatchulidwa pamwambapa, zomwe zikuwonetsa mawonekedwe osalekeza.16S rRNA amplicon sequencing adapanga 29,285 motsatira, kuyimira mitundu 346.Kuphatikizana ndi mbiri ya zamoyo pogwiritsa ntchito ma amplicon sequencing, kufalitsa kumeneku kunapezeka kuti sikunachokere ku microbiota yokhudzana ndi mizu, komabe, ikhoza kuthandizidwa ndi mafoni amtundu wazinthu.Kafukufukuyu adazindikira kutuluka kwa ma ARGs ndi tizilombo toyambitsa matenda kuchokera m'nthaka kulowa muzomera kudzera munjira yolumikizana ndi mizu ya nthaka.

Buku

Wang, C. , Hu, R. , Strong, PJ , Zhuang, W. , & Shu, L. .(2020).Kuchuluka kwa majini olimbana ndi maantibayotiki ndi tizilombo toyambitsa matenda m'nthaka-mizu ya mangrove mosalekeza.Journal of Hazardous Materials, 408, 124985.