304 Stainless hlau welded coiled raj / tubing zhemical zomponent, Biosynthetic muaj peev xwm ntawm lub ntiaj teb no Marine Microbiome

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Sliders qhia peb kab lus rau ib tus swb.Siv cov nyees khawm rov qab thiab tom ntej kom txav mus los ntawm cov slides, lossis cov khawm tswj swb thaum kawg kom txav mus los ntawm txhua tus swb.

Cov khoom piav qhia ntxaws

304 Stainless hlau welded coiled raj / tubing
1. Specification: Stainless hlau coil tubing / tubing
2. Hom: welded los yog seamless
3. Txuj: ASTM A269, ASTM A249
4. Stainless hlau coil tube OD: 6mm txog 25.4MM
5. Ntev: 600-3500MM lossis raws li tus neeg siv khoom xav tau.
6. Phab ntsa thickness: 0.2mm txog 2.0mm.

7. Kev kam rau siab: OD: +/- 0.01mm;Thickness: +/- 0.01%.

8. Coil puab qhov loj: 500MM-1500MM (tuaj yeem kho raws li cov neeg siv khoom xav tau)

9. Coil qhov siab: 200MM-400MM (tuaj yeem kho raws li cov neeg siv khoom xav tau)

10. Nto: Ci los yog annealed
11. Khoom siv: 304, 304L, 316L, 321, 301, 201, 202, 409, 430, 410, alloy 625, 825, 2205, 2507, thiab lwm yam.
12. Packing: woven hnab nyob rau hauv cov ntaub ntawv ntoo, ntoo pallet, ntoo ncej, los yog raws li ib tus neeg yuav tsum tau muaj
13. Test: tshuaj tivthaiv, yield zog, tensile zog, hardness ntsuas
14. Guarantee: Tus neeg thib peb (piv txwv li: SGS TV) tshuaj xyuas, thiab lwm yam.
15. Daim ntawv thov: Kho kom zoo nkauj, rooj tog, roj tsheb thauj mus los, tshav kub exchanger, railing ua, ntawv ua, tsheb, khoom noj khoom haus, kev kho mob, thiab lwm yam.

Ntuj microbial zej zog yog phylogenetically thiab metabolically ntau haiv neeg.Ntxiv nrog rau cov pab pawg tsis tau kawm ntawm cov kab mob 1, qhov kev sib txawv no kuj muaj peev xwm ua kom muaj txiaj ntsig zoo rau kev tshawb pom ntawm ecologically thiab biotechnologically tseem ceeb enzymes thiab biochemical compounds2,3.Txawm li cas los xij, kev kawm txog kev sib txawv no los txiav txim siab txog txoj hauv kev genomic uas sib txuas cov khoom sib txuas thiab khi lawv rau lawv cov tswv cuab tseem yog qhov nyuaj.Lub peev xwm biosynthetic ntawm cov kab mob hauv dej hiav txwv qhib tseem tsis tau paub ntau vim muaj kev txwv hauv kev txheeb xyuas tag nrho cov ntaub ntawv daws teeb meem genome ntawm lub ntiaj teb.Ntawm no, peb tshawb nrhiav qhov sib txawv thiab ntau haiv neeg ntawm biosynthetic gene pawg hauv dej hiav txwv los ntawm kev sib koom ua ke txog 10,000 microbial genomes los ntawm cov kab lis kev cai thiab ib lub hlwb nrog ntau dua 25,000 tus qauv tsim tshiab tshiab los ntawm ntau dua 1,000 cov qauv dej hiav txwv.Cov kev siv zog no tau txheeb xyuas txog 40,000 putative feem ntau tshiab biosynthetic gene pawg, qee qhov tau pom nyob rau hauv pawg phylogenetic yav dhau los uas tsis xav txog.Hauv cov pej xeem no, peb tau txheeb xyuas cov kab mob uas muaj nyob hauv biosynthetic gene pawg ("Candidatus Eudormicrobiaceae") uas yog cov kab mob uas tsis muaj kab mob thiab suav nrog qee cov kab mob biosynthetically ntau haiv neeg nyob hauv ib puag ncig no.Ntawm cov no, peb tau ua tus yam ntxwv ntawm phosphatase-peptide thiab pytonamide txoj hauv kev, txheeb xyuas cov xwm txheej ntawm cov qauv sib txawv bioactive compound thiab enzymology, feem.Hauv kev xaus, txoj kev tshawb no qhia tau hais tias yuav ua li cas microbiome-raws li cov tswv yim tuaj yeem pab tshawb nrhiav cov enzymes yav dhau los uas tsis tau piav qhia thiab cov khoom noj ntuj nyob hauv qhov tsis to taub microbiota thiab ib puag ncig.
Microbes tsav lub ntiaj teb biogeochemical cycles, tswj cov zaub mov webs, thiab ua kom cov nroj tsuag thiab tsiaj noj qab haus huv5.Lawv loj heev phylogenetic, metabolic thiab functional ntau haiv neeg sawv cev rau ib tug nplua nuj muaj peev xwm rau nrhiav tau ntawm tshiab taxa1, enzymes thiab biochemical compounds, nrog rau tej yam ntuj tso khoom6.Hauv cov zej zog ecological, cov molecules no muab cov kab mob sib kis nrog ntau yam kev ua haujlwm ntawm lub cev thiab ecological, los ntawm kev sib txuas lus mus rau kev sib tw 2, 7.Ntxiv rau lawv cov haujlwm qub, cov khoom siv ntuj tsim no thiab lawv txoj hauv kev tsim cov noob caj noob ces muab piv txwv rau kev siv biotechnological thiab kho mob 2,3.Kev txheeb xyuas ntawm txoj hauv kev thiab kev sib txuas tau zoo heev los ntawm kev kawm txog kab lis kev cai microbes.Txawm li cas los xij, kev tshawb fawb taxonomic ntawm ib puag ncig ntuj tau pom tias feem coob ntawm cov kab mob tsis tau cog qoob loo8.Qhov kev coj noj coj ua no txwv peb lub peev xwm los siv cov kev ua haujlwm sib txawv uas tau muab zais los ntawm ntau cov kab mob microbes4,9.
Txhawm rau kov yeej cov kev txwv no, kev txhim kho thev naus laus zis hauv kaum xyoo dhau los tau tso cai rau cov kws tshawb fawb ncaj qha (piv txwv li, tsis muaj kab lis kev cai ua ntej) cov kab ke DNA microbial los ntawm tag nrho cov zej zog (metagenomics) lossis tib lub hlwb.Lub peev xwm los sib sau ua ke cov fragments no rau hauv cov genome loj dua thiab rov tsim kho ntau yam metagenomically sib sau ua ke genomes (MAGs) lossis ib qho amplified genomes (SAGs), feem, qhib lub sijhawm tseem ceeb rau kev tshawb fawb taxocentric ntawm microbiome (ie, cov zej zog microbial thiab microbiome).taug txoj kev tshiab.tus kheej cov khoom siv caj ces hauv ib puag ncig muab) 10,11,12.Tseeb tiag, cov kev tshawb fawb tsis ntev los no tau nthuav dav ntau qhov phylogenetic sawv cev ntawm microbial ntau haiv neeg hauv ntiaj teb 1, 13 thiab tau nthuav tawm ntau yam kev ua haujlwm sib txawv hauv cov zej zog microbial uas tsis tau hais los ntawm kab lis kev cai kab lis kev cai siv genome sequences (REFs) 14.Lub peev xwm los tso qhov tsis pom kev ua haujlwm sib txawv hauv cov ntsiab lus ntawm tus tswv tsev genome (ie, genome daws teeb meem) yog qhov tseem ceeb rau kev kwv yees li tseem tsis tau paub txog cov kab mob microbial uas xav tias yuav nkag mus rau cov khoom ntuj tshiab 15,16 lossis rau kev taug qab cov khoom sib txuas rov qab mus rau lawv cov thawj tsim17.Piv txwv li, ib qho kev sib koom ua ke ntawm metagenomic thiab ib leeg-cell genomic tsom xam txoj hauv kev tau coj mus rau kev txheeb xyuas ntawm Candidatus Entotheonella, ib pawg ntawm cov kab mob metabolically nplua nuj sponge-associated bacteria, raws li cov neeg tsim khoom ntawm ntau yam tshuaj muaj peev xwm18.Txawm li cas los xij, txawm tias tsis ntev los no tau sim ntawm kev tshawb nrhiav genomic ntawm ntau haiv neeg microbial, 16,19 ntau dua li ob feem peb ntawm cov ntaub ntawv thoob ntiaj teb metagenomic rau lub ntiaj teb cov dej hiav txwv loj tshaj plaws ntawm ecosystems16,20 tseem ploj lawm.Yog li, feem ntau, biosynthetic muaj peev xwm ntawm marine microbiome thiab nws lub peev xwm ua ib qho chaw cia khoom ntawm cov khoom tshiab enzymatic thiab cov khoom ntuj tsim nyob rau hauv ntau understudied.
Txhawm rau tshawb nrhiav lub peev xwm biosynthetic ntawm marine microbiomes nyob rau hauv lub ntiaj teb no, peb thawj zaug sib sau ua ke marine microbial genomes tau siv kab lis kev cai thiab tsis yog kab lis kev cai los tsim cov ntaub ntawv dav dav ntawm phylogenetics thiab noob muaj nuj nqi.Kev tshuaj xyuas ntawm cov ntaub ntawv no tau qhia ntau yam ntawm biosynthetic gene pawg (BGCs), feem ntau ntawm cov tsev neeg uas tseem tsis tau paub txog cov noob caj noob ces (GCF).Tsis tas li ntawd, peb tau txheeb xyuas ib tsev neeg tsis paub kab mob uas nthuav tawm qhov paub ntau tshaj plaws ntawm BGCs hauv dej hiav txwv qhib rau hnub tim.Peb xaiv ob txoj kev sib txuas ntawm ribosomal thiab post-translationally modified peptide (RiPP) txoj hauv kev rau kev sim validation raws li lawv cov caj ces sib txawv ntawm txoj kev paub tam sim no.Kev ua haujlwm zoo ntawm txoj hauv kev no tau nthuav tawm cov piv txwv tsis xav txog ntawm enzymology nrog rau cov khoom sib txawv txawv txawv nrog kev ua haujlwm protease inhibitory.
Thaum xub thawj, peb tsom los tsim cov ntaub ntawv thoob ntiaj teb rau kev tshuaj ntsuam genome, tsom mus rau nws cov kab mob thiab cov archaeal.Txog rau qhov kawg no, peb tau sau cov ntaub ntawv metagenomic thiab 1038 cov qauv dej hiav txwv los ntawm 215 qhov chaw kuaj thoob ntiaj teb (latitude range = 141.6°) thiab ob peb txheej sib sib zog nqus (los ntawm 1 txog 5600 m hauv qhov tob, npog thaj tsam pelagic, mesopelagic thiab abyssal).Keeb kwm 21,22,23 (Daim duab 1a, cov ntaub ntawv txuas ntxiv, Daim duab 1a thiab Cov Lus Qhia Ntxiv 1).Ntxiv nrog rau kev muab thaj chaw dav dav, cov kev xaiv lim dej no tso cai rau peb los sib piv ntau yam ntawm marine microbiome, suav nrog cov kab mob nplua nuj (<0.2 µm), prokaryotic-nplua nuj (0.2–3 µm), particle-nplua nuj (0.8 µm). ).-20 µm) thiab tus kab mob-depleted (> 0.2 µm) colony.
a, Tag nrho ntawm 1038 cov genomes uas muaj nyob rau hauv pej xeem (metagenomics) ntawm marine microbial cov zej zog sau los ntawm 215 qhov chaw thoob ntiaj teb (62 ° S txog 79 ° N thiab 179 ° W txog 179 ° E .).Daim ntawv qhia pobzeb © Esri.Qhov chaw: GEBCO, NOAA, CHS, OSU, UNH, CSUMB, National Geographic, DeLorme, NAVTEQ, thiab Esri.b, cov metagenomes no tau siv los tsim kho MAGs (txoj kev thiab cov ntaub ntawv ntxiv), uas txawv ntawm qhov ntau thiab zoo (txoj kev) hauv cov ntaub ntawv (cim xim).Cov MAGs rov tsim kho tau ntxiv nrog rau pej xeem (sab nraud) genomes, suav nrog handcrafted MAG26, SAG27 thiab REF.27 Compile OMD.c, piv rau yav dhau los cov ntaub ntawv raws li tsuas yog SAG (GORG)20 lossis MAG (GEM)16, OMD txhim kho genomic characterization ntawm marine microbial zej zog (metagenomic nyeem daim ntawv qhia tus nqi; txoj kev) los ntawm ob mus rau peb zaug nrog ntau tus sawv cev nyob rau hauv qhov tob thiab latitude..<0.2, n=151, 0.2-0.8, n=67, 0.2-3, n=180, 0.8-20, n=30, >0.2, n=610, <30°, n=132, 30–60° , n = 73, > 60°, n = 42, EPI, n = 174, MES, n = 45, BAT, n = 28. d, OMD pab pawg ua hom pawg (95% txhais tau tias nucleotide tus kheej) txheeb xyuas tag nrho cov kwv yees li 8300 hom, ntau tshaj li ib nrab ntawm cov uas tsis tau ua dhau los ua tus yam ntxwv raws li kev sau se ntawm cov ntaub ntawv siv GTDB (version 89) e, kev faib tawm ntawm hom los ntawm hom genome tau pom tias MAG, SAG thiab REFs sib koom ua ke zoo hauv kev xav txog kev sib txawv ntawm phylogenetic. marine microbiome.Hauv particular, 55%, 26% thiab 11% ntawm hom yog tshwj xeeb rau MAG, SAG thiab REF, raws li.BATS, Bermuda Atlantic Sijhawm Series;GEM, genomes ntawm lub ntiaj teb microbiome;GORG, thoob ntiaj teb dej hiav txwv siv genome;HOT, Hawaiian Dej hiav txwv lub sij hawm series.
Siv cov ntaub ntawv no, peb rov tsim dua tag nrho 26,293 MAGs, feem ntau yog kab mob thiab archaeal (Fig. 1b thiab nthuav cov ntaub ntawv, Fig. 1b).Peb tsim cov MAGs los ntawm cov rooj sib txoos los ntawm kev sib cais es tsis yog cov qauv sib xyaw ua ke los tiv thaiv kev tawg ntawm qhov sib txawv ntawm cov qauv ntawm cov qauv los ntawm qhov chaw sib txawv lossis lub sijhawm (txoj kev).Tsis tas li ntawd, peb tau muab cov genomic fragments raws li lawv cov kev sib raug zoo nyob thoob plaws ntau tus qauv (los ntawm 58 mus rau 610 tus qauv, nyob ntawm kev soj ntsuam; txoj kev).Peb pom tias qhov no yog lub sijhawm siv sijhawm, tab sis tseem ceeb step24 uas tau hla hauv ntau qhov loj MAG16, 19, 25 rov tsim kho thiab txhim kho kom muaj nuj nqis (2.7-fold ntawm qhov nruab nrab) thiab zoo (+ 20% ntawm qhov nruab nrab) ntawm cov genome.reconstructed los ntawm marine metagenome kawm ntawm no (cov ntaub ntawv txuas ntxiv, Fig. 2a thiab cov ntaub ntawv ntxiv).Zuag qhia tag nrho, cov kev siv zog no tau ua rau 4.5-fold nce hauv marine microbial MAGs (6-fold yog tias tsuas yog MAGs zoo raug suav hais tias) piv rau cov peev txheej MAG uas muaj nyob rau niaj hnub no16 (Txoj Kev).Cov txheej txheem MAG tshiab no tau ua ke nrog 830 tes xaiv MAG26s, 5969 SAG27s thiab 1707 REFs.Nees nkaum xya hom kab mob marine thiab archaea ua ib qho kev sib sau ua ke ntawm 34,799 genomes (Fig. 1b).
Tom qab ntawd peb tau soj ntsuam cov peev txheej tsim tshiab los txhim kho nws lub peev xwm los sawv cev rau cov zej zog marine microbial thiab ntsuas qhov cuam tshuam ntawm kev sib koom ua ke ntau hom genome.Qhov nruab nrab, peb pom tias nws npog kwv yees li 40-60% ntawm cov ntaub ntawv marine metagenomic (Daim duab 1c), ob mus rau peb npaug ntawm cov ntaub ntawv tshaj tawm ntawm MAG-tsuas yog nyob rau hauv ob qho tib si qhov tob thiab latitude Ntau serial 16 lossis SAG20.Tsis tas li ntawd, txhawm rau ntsuas qhov sib txawv ntawm kev sau se hauv cov ntawv sau, peb sau tseg tag nrho cov genomes siv Genome Taxonomy Database (GTDB) cov cuab yeej (txoj kev) thiab siv qhov nruab nrab genome-wide nucleotide tus kheej ntawm 95%.28 txhawm rau txheeb xyuas 8,304 hom pawg (npe).Ob feem peb ntawm cov hom no (nrog rau cov clades tshiab) tsis tau tshwm sim yav dhau los hauv GTDB, ntawm 2790 tau tshawb pom siv MAG reconstructed hauv txoj kev tshawb no (Fig. 1d).Tsis tas li ntawd, peb pom tias ntau hom genomes muaj ntau qhov sib ntxiv: 55%, 26%, thiab 11% ntawm hom yog tsim los ntawm MAG, SAG, thiab REF, raws li (Fig. 1e).Tsis tas li ntawd, MAG tau npog tag nrho 49 hom pom nyob rau hauv kab dej, thaum SAG thiab REF tsuas yog sawv cev rau 18 thiab 11 ntawm lawv, feem.Txawm li cas los xij, SAG zoo dua sawv cev rau ntau haiv neeg ntawm cov clades feem ntau (cov ntaub ntawv nthuav dav, Fig. 3a), xws li Pelagic Bacteriales (SAR11), nrog SAG npog yuav luag 1300 hom thiab MAG tsuas yog 390 hom.Qhov tseem ceeb, REFs tsis tshua sib tshooj nrog MAGs lossis SAGs ntawm hom kab thiab sawv cev> 95% ntawm kwv yees li 1000 genomes tsis pom nyob rau hauv cov dej hiav txwv qhib metagenomic tau kawm ntawm no, feem ntau yog vim muaj kev cuam tshuam nrog lwm hom kev sib cais ntawm cov neeg sawv cev marine (xws li sediments ) .los yog host-associate).Txhawm rau ua kom muaj dav dav rau cov zej zog kev tshawb fawb, cov peev txheej marine genome, uas tseem suav nrog cov khoom tsis sib xws (piv txwv li, los ntawm kev kwv yees phages, genomic Islands, thiab genome fragments uas muaj cov ntaub ntawv tsis txaus rau MAG rov tsim dua), tuaj yeem muab piv nrog cov ntaub ntawv taxonomic. .Nkag mus rau cov lus piav qhia nrog rau cov noob muaj nuj nqi thiab cov ntsiab lus tsis sib xws hauv Ocean Microbiology Database (OMD; https://microbiomics.io/ocean/).
Tom qab ntawd peb npaj tawm los tshawb txog kev nplua nuj thiab tshiab ntawm biosynthetic muaj peev xwm hauv dej hiav txwv qhib microbiomes.Txog rau qhov kawg no, peb thawj zaug siv tshuaj tiv thaiv SMASH rau tag nrho MAGs, SAGs, thiab REFs pom nyob rau hauv 1038 marine metagenomes (txoj kev) los kwv yees tag nrho ntawm 39,055 BGCs.Tom qab ntawd peb muab cov no tso rau hauv 6907 GCFs uas tsis yog rov ua dua thiab 151 gene pawg pawg neeg (GCCs; Ntxiv Table 2 thiab txoj hauv kev) kom suav rau qhov rov ua dua tshiab (piv txwv li, tib BGC tuaj yeem nkag mus rau hauv ntau lub genomes) thiab cov ntaub ntawv metagenomic Fragmentation ntawm concentrated BGCs .BGCs tsis tiav tsis tau nce ntxiv, yog tias muaj (Cov Lus Qhia Ntxiv), tus naj npawb ntawm GCFs thiab GCCs, raws li, muaj tsawg kawg yog ib tus tswvcuab BGC tsis zoo hauv 44% thiab 86% ntawm cov neeg mob.
Nyob rau theem GCC, peb pom ntau yam kev kwv yees RiPPs thiab lwm yam khoom ntuj tsim (Fig. 2a).Ntawm lawv, piv txwv li, arylpolyenes, carotenoids, ectoines, thiab siderophores yog GCCs nrog kev faib tawm dav dav thiab muaj kev nplua nuj nyob hauv dej hiav txwv metagenomes, uas yuav qhia tau tias muaj kev hloov pauv ntawm cov kab mob mus rau hauv ib puag ncig marine, suav nrog kev ua haujlwm rau cov pa oxygen reactive, oxidative thiab osmotic stress..los yog hlau nqus (ntau ntxiv).Qhov kev ua haujlwm sib txawv no sib txawv nrog kev tshuaj xyuas tsis ntev los no ntawm kwv yees li 1.2 lab BGCs ntawm kwv yees li 190,000 genomes khaws cia hauv NCBI RefSeq database (BiG-FAM/RefSeq, tom qab ntawd hu ua RefSeq)29, uas pom tias nonribosomal Synthetase peptides thiab polyketides (NRPS) (PKS) BGCs (Cov Ntaub Ntawv Ntxiv).Peb kuj pom 44 (29%) GCCs tsuas yog nyob deb ntawm txhua qhov RefSeq BGC (\(\bar{d}\)RefSeq> 0.4; Fig. 2a thiab txoj kev) thiab 53 (35%) GCCs nkaus xwb hauv MAG , qhia txog lub peev xwm txhawm rau txheeb xyuas cov tshuaj tsis tau piav qhia yav dhau los hauv OMD.Muab hais tias txhua tus GCCs no yuav sawv cev rau ntau hom kev ua haujlwm biosynthetic, peb tau txheeb xyuas cov ntaub ntawv ntxiv ntawm GCF qib hauv kev sib zog los muab cov ncauj lus kom ntxaws ntxiv ntawm BGCs kwv yees rau code rau cov khoom zoo sib xws29.Tag nrho ntawm 3861 (56%) tau txheeb xyuas GCFs tsis sib tshooj nrog RefSeq, thiab> 97% ntawm GCFs tsis muaj nyob hauv MIBiG, yog ib qho ntawm cov ntaub ntawv loj tshaj plaws ntawm kev sim BGCs (Daim duab 2b).Txawm hais tias nws tsis yog qhov xav tsis thoob los nrhiav pom ntau txoj hauv kev tshiab hauv cov chaw uas tsis zoo sawv cev los ntawm kev siv genome, peb txoj hauv kev rau dereplicating BGCs rau hauv GCFs ua ntej benchmarking txawv los ntawm yav dhau los qhia 16 thiab tso cai rau peb muab ib qho kev ntsuam xyuas tsis ncaj ncees lawm ntawm novelty.Feem ntau ntawm ntau haiv neeg tshiab (3012 GCF los yog 78%) sib raug rau terpenes, RiPP lossis lwm yam khoom siv ntuj tsim, thiab feem ntau (1815 GCF lossis 47%) yog encoded yam tsis paub vim lawv lub peev xwm biosynthetic.Tsis zoo li PKS thiab NRPS pawg, cov BGCs compact no tsis tshua muaj feem cuam tshuam thaum lub sij hawm sib dhos metagenomic 31 thiab tso cai rau ntau lub sij hawm- thiab cov peev txheej ua haujlwm zoo ntawm lawv cov khoom.
Tag nrho ntawm 39,055 BGCs tau muab faib ua 6,907 GCFs thiab 151 GCCs.a, cov ntaub ntawv sawv cev (internal sab nraud).Hierarchical pawg ntawm BGC nrug raws li GCC, 53 ntawm uas yog kho los ntawm MAG nkaus xwb.GCC muaj BGCs los ntawm txawv taxa (ln-transformed rooj vag zaus) thiab txawv BGC chav kawm (lub voj voog loj sib haum rau nws zaus).Rau txhua GCC, txheej txheej sab nrauv sawv cev rau tus naj npawb ntawm BGCs, qhov tshwm sim (feem pua ​​​​ntawm cov qauv), thiab qhov kev ncua deb (tsawg kawg BGC cosine nrug (min(dMIBiG))) los ntawm BiG-FAM mus rau BGC.GCCs nrog BGCs ze ze rau kev sim tshuaj ntsuam xyuas BGCs (MIBiG) yog qhov tseem ceeb nrog cov xub.b Sib piv GCF nrog kev kwv yees (BiG-FAM) thiab kev sim ntsuas (MIBiG) BGCs, 3861 tshiab (d–> 0.2) GCFs tau pom.Feem ntau (78%) ntawm cov cai no rau RiPP, terpenes, thiab lwm yam khoom siv ntuj tsim.c, tag nrho cov genomes hauv OMD pom nyob rau hauv 1038 marine metagenomes tau muab tso rau hauv GTDB lub hauv paus tsob ntoo los qhia txog phylogenetic kev pab them nqi ntawm OMD.Clades tsis muaj genomes hauv OMD yog pom hauv grey.Tus naj npawb ntawm BGCs sib raug rau tus lej loj tshaj plaws ntawm kev kwv yees BGCs ib lub genome hauv ib daim ntawv muab.Txhawm rau kom pom tseeb, 15% kawg ntawm cov nodes tau tawg.Cov xub qhia clades nplua nuj nyob rau hauv BGC (> 15 BGC), tshwj tsis yog Mycobacterium, Gordonia (thib ob rau Rhodococcus), thiab Crocosphaera (thib ob rau Synechococcus).d, npe c.Eremiobacterota tau pom qhov ntau tshaj plaws biosynthetic ntau haiv neeg (Shannon Performance index raws li hom khoom ntuj).Txhua pawg sawv cev rau genome nrog BGCs feem ntau hauv hom.T1PKS, PKS hom I, T2/3PKS, PKS hom II thiab hom III.
Ntxiv nrog rau kev nplua nuj thiab tshiab, peb tshawb nrhiav cov qauv biogeographic ntawm biosynthetic peev xwm ntawm marine microbiome.Kev sib faib ua piv txwv los ntawm qhov nruab nrab metagenomic GCF daim ntawv luam tus lej faib (Txoj Kev) tau pom tias qis-latitude, nto, prokaryotic-nplua nuj thiab cov kab mob-cov zej zog tsis zoo, feem ntau los ntawm cov dej saum npoo av los yog tob hnub ci, muaj nplua nuj nyob hauv RiPP thiab BGC terpenes.Hauv qhov sib piv, lub ncov qaumteb qabteb, dej hiav txwv tob, kab mob- thiab cov zej zog nplua nuj muaj feem cuam tshuam nrog ntau dua ntawm NRPS thiab PKS BGC (cov ntaub ntawv nthuav dav, daim duab 4 thiab cov ntaub ntawv ntxiv).Thaum kawg, peb pom tias cov zej zog uas tau kawm zoo thiab cov pelagic yog qhov muaj txiaj ntsig zoo tshaj plaws ntawm cov terpenes tshiab (Augmented Data Figure).Lub peev xwm siab tshaj plaws rau PKS, RiPP thiab lwm yam khoom ntuj tsim (Daim duab 5a nrog cov ntaub ntawv nthuav dav).
Txhawm rau ua kom tiav peb txoj kev tshawb fawb txog biosynthetic muaj peev xwm ntawm marine microbiomes, peb tsom los qhia lawv cov kev faib tawm phylogenetic thiab txheeb xyuas cov BGC-enriched clades tshiab.Txog rau qhov kawg no, peb muab cov genomes ntawm marine microbes tso rau hauv cov kab mob zoo li qub GTDB13 cov kab mob thiab archaeal phylogenetic ntoo thiab overlaid lub putative biosynthetic pathways lawv encode (Fig. 2c).Peb tau yooj yim kuaj pom ob peb BGC-enriched clades ( sawv cev los ntawm ntau dua 15 BGCs) hauv cov qauv dej hiav txwv (txoj kev) paub txog lawv cov peev xwm biosynthetic, xws li cyanobacteria (Synechococcus) thiab cov kab mob Proteus, xws li Tistrella32,33, lossis tsis ntev los no tau txais kev nyiam rau lawv. natural khoom.xws li Myxococcota (Sandaracinaceae), Rhodococcus thiab Planctomycetota34,35,36.Interestingly, peb pom ob peb yav dhau los tsis tau tshawb nrhiav kab hauv cov kab no.Piv txwv li, cov tsiaj uas muaj peev xwm biosynthetic nplua nuj nyob rau hauv phyla Planctomycetota thiab Myxococcota tau koom nrog cov neeg sib tw tsis sib xws thiab cov genera, ntsig txog (Table 3).Ua ke, qhov no qhia tias OMD muab kev nkag mus rau yav dhau los tsis paub txog cov ntaub ntawv phylogenetic, suav nrog cov kab mob, uas tuaj yeem sawv cev rau lub hom phiaj tshiab rau enzyme thiab kev tshawb pom cov khoom ntuj.
Tom ntej no, peb qhia txog BGC-enriched clade los ntawm tsis tsuas yog suav tus lej siab tshaj plaws ntawm BGCs encoded los ntawm nws cov tswv cuab, tab sis kuj los ntawm kev ntsuam xyuas qhov sib txawv ntawm cov BGCs, uas piav qhia qhov zaus ntawm ntau hom khoom sib txawv ntawm ntuj tsim (Daim duab 2c thiab cov txheej txheem )..Peb pom tias ntau hom biosynthetically ntau haiv neeg tau sawv cev los ntawm cov kab mob tshwj xeeb tsim MAGs hauv txoj kev tshawb no.Cov kab mob no muaj nyob rau hauv uncultivated phylum Candidatus Eremiobacterota, uas tseem unexplored loj sib nrug los ntawm ob peb genomic kev tshawb fawb37,38.Nws yog noteworthy tias "ca.Lub genus Eremiobacterota tsuas yog tau txheeb xyuas hauv thaj chaw thaj av39 thiab tsis paub tias suav nrog cov tswvcuab ntxiv rau BGC.Ntawm no peb tau rov tsim dua yim MAGs ntawm tib hom (nucleotide tus kheej> 99%) 23. Yog li ntawd peb thiaj li tawm tswv yim hom npe "Candidatus Eudoremicrobium malaspinii", uas muaj npe tom qab nereid (hiav txwv nymph), khoom plig zoo nkauj hauv Greek mythology thiab ntoj ke mus kawm.'Ka.Raws li phylogenetic annotation 13, E. malaspinii tsis muaj cov txheeb ze paub yav dhau los hauv qab theem sib lawv liag thiab yog li ntawd yog tsev neeg kab mob tshiab uas peb hais tias "Ca.E. malaspinii” raws li hom hom thiab “Ca.Eudormicrobiaceae" raws li lub npe (Cov Ntaub Ntawv Ntxiv).Luv luv metagenomic reconstruction ntawm 'Ca.E. malaspinii genome project tau raug lees paub los ntawm cov tswv yim tsawg heev, nyeem ntev metagenomic sequencing thiab phiaj los ua ke ntawm ib qho qauv (Txoj Kev) raws li ib qho 9.63 Mb linear chromosome nrog 75 kb duplication.raws li qhov tseem tshuav tsis meej pem.
Txhawm rau tsim kom muaj cov ntsiab lus phylogenetic ntawm cov tsiaj no, peb tau tshawb nrhiav 40 hom kab mob sib xws hauv cov qauv eukaryotic-enriched metagenomic ntxiv los ntawm Tara Dej hiav txwv ntoj ke mus los ntawm kev tsim kho genome.Luv luv, peb tau txuas metagenomic nyeem rau genomic fragments txuam nrog "Ca.E. malaspinii” thiab xav tias kev nrhiav neeg ua haujlwm nce ntxiv hauv cov qauv no qhia tias muaj lwm cov txheeb ze (txoj kev).Raws li qhov tshwm sim, peb pom 10 MAGs, kev sib xyaw ntawm 19 MAGs sawv cev rau tsib hom hauv peb genera hauv ib tsev neeg tshiab (ie "Ca. Eudormicrobiaceae").Tom qab kev tshuaj xyuas phau ntawv thiab kev tswj kom zoo (cov ntaub ntawv nthuav dav, daim duab 6 thiab cov ntaub ntawv ntxiv), peb pom tias “Ca.Eudormicrobiaceae hom muaj cov genomes loj dua (8 Mb) thiab muaj peev xwm ntau dua biosynthetic (14 txog 22 BGC ib hom) dua li lwm cov tswv cuab "Ca".Clade Eremiobacterota (txog 7 BGC) (Fig. 3a–c).
a, Phylogogenetic txoj hauj lwm ntawm tsib 'Ca.Hom Eudormicrobiaceae tau pom BGC nplua nuj tshwj xeeb rau cov kab hauv hiav txwv tau txheeb xyuas hauv txoj kev tshawb no.Cov ntoo phylogenetic suav nrog tag nrho 'Ca.MAG Eremiobacterota thiab cov tswv cuab ntawm lwm cov phyla (genome tooj nyob rau hauv brackets) muab nyob rau hauv GTDB (version 89) tau siv rau evolutionary keeb kwm yav dhau (Txoj kev).Cov khaubncaws sab nraud povtseg sawv cev rau kev faib tawm ntawm tsev neeg (“Ca. Eudormicrobiaceae” thiab “Ca. Xenobiaceae”) thiab hauv chav kawm (“Ca. Eremiobacteria”).Cov tsib hom uas tau piav qhia hauv txoj kev tshawb no yog sawv cev los ntawm cov lej lej thiab cov npe binomial (Cov Lus Qhia Ntxiv).b ,ua ok.Eudormicrobiaceae hom muaj xya hom BGC nuclei.Qhov tsis tuaj ntawm BGC hauv A2 clade yog vim qhov ua tsis tiav ntawm tus neeg sawv cev MAG (Table Ntxiv 3).BGCs tshwj xeeb rau “Ca.Amphithomicrobium" thiab "Ca.Amphithomicrobium" (clades A thiab B) tsis qhia.c, Txhua BGCs encoded li “Ca.Eudoremicrobium taraoceanii tau pom tau hais tawm hauv 623 metatranscriptomes coj los ntawm cov dej hiav txwv ntawm Tara.Cov voj voog khoom qhia tias muaj kev hloov pauv.Lub voj voog txiv kab ntxwv qhia txog lub log2-hloov pauv hloov pauv hauv qab thiab saum toj no ntawm kev tu vaj tse hauv tsev (txoj kev).d, txheeb ze abundance curves (txoj kev) qhia 'Ca.Hom Eudormicrobiaceae yog dav nyob rau hauv feem ntau ntawm cov dej hiav txwv bass thiab nyob rau hauv tag nrho cov dej kem (los ntawm qhov chaw mus rau ib tug tob ntawm tsawg kawg yog 4000 m).Raws li cov kev kwv yees no, peb pom tias 'Ca.E. malaspinii 'sau txog li 6% ntawm cov hlwb prokaryotic nyob rau hauv dej hiav txwv pelagic nplej-txuas nrog cov zej zog.Peb txiav txim siab ib hom yuav tsum muaj nyob rau ntawm ib qhov chaw yog tias nws pom nyob rau hauv ib feem ntawm qhov loj ntawm ib tug muab qhov tob txheej.IO – Indian Ocean, NAO – North Atlantic, NPO – North Pacific, RS – Red Hiav txwv, SAO – South Atlantic, SO – Southern Ocean, SPO – South Pacific.
Kawm txog kev nplua nuj thiab kev faib tawm ntawm Ca.Eudormicrobiaceae, uas, raws li peb pom, predominates nyob rau hauv feem ntau dej hiav txwv basins, raws li zoo raws li nyob rau hauv tag nrho cov dej kem (Fig. 3d).Hauv zos, lawv suav txog 6% ntawm marine microbial zej zog, ua rau lawv yog ib feem tseem ceeb ntawm lub ntiaj teb marine microbiome.Tsis tas li ntawd, peb pom cov ntsiab lus txheeb ze ntawm Ca.Eudormicrobiaceae hom thiab lawv cov BGC qhia theem siab tshaj plaws nyob rau hauv eukaryotic enriched feem (Fig. 3c thiab txuas ntxiv cov ntaub ntawv, Fig. 7), qhia tau hais tias muaj kev cuam tshuam nrog cov khoom me me, nrog rau cov plankton.Qhov kev soj ntsuam no dais qee qhov zoo ib yam li 'Ca.Eudoremicrobium BGCs uas tsim cov khoom lag luam cytotoxic los ntawm txoj hauv kev paub yuav ua rau pom tus cwj pwm predatory (Cov ntaub ntawv ntxiv thiab nthuav cov ntaub ntawv, daim duab 8), zoo ib yam li lwm cov tsiaj txhu uas tsim cov metabolites xws li Myxococcus41.Discovery ntawm Ca.Eudormicrobiaceae nyob rau hauv tsawg muaj (dej hiav txwv tob) los yog eukaryotic es tsis yog cov qauv prokaryotic yuav piav qhia yog vim li cas cov kab mob no thiab lawv cov kev npaj txhij txog BGC ntau haiv neeg tseem tsis meej nyob rau hauv cov ntsiab lus ntawm kev tshawb fawb zaub mov.
Thaum kawg, peb nrhiav kev sim ua pov thawj cov lus cog tseg ntawm peb txoj haujlwm microbiome hauv kev tshawb nrhiav txoj hauv kev tshiab, enzymes, thiab cov khoom ntuj tsim.Ntawm cov chav kawm sib txawv ntawm BGCs, RiPP txoj hauv kev paub txog kev nkag siab cov tshuaj lom neeg thiab kev ua haujlwm ntau yam vim muaj ntau yam kev hloov pauv tom qab hloov pauv ntawm cov tub ntxhais peptide los ntawm cov enzymes paub tab42.Yog li ntawd peb xaiv ob 'Ca.Eudoremicrobium 'RiPP BGCs (Figures 3b thiab 4a-e) yog ua raws tib yam li ib qho BGC paub (\(\bar{d}\)MIBiG thiab \(\bar{d}\)RefSeq saum 0.2).
a–c, In vitro heterologous expression thiab in vitro enzymatic assays of a novel (\(\bar{d}\)RefSeq = 0.29) pawg ntawm RiPP biosynthesis tshwj xeeb rau cov tsiaj hiav txwv sib sib zog nqus Ca.E. malaspinii' coj mus rau kev tsim cov khoom lag luam diphosphorylated.c, kev hloov kho uas siv cov kev daws teeb meem siab (HR) MS/MS (fragmentation qhia los ntawm b thiab y ions hauv cov qauv tshuaj) thiab NMR (cov ntaub ntawv nthuav dav, daim duab 9).d, phosphorylated peptide no nthuav tawm tsawg micromolar inhibition ntawm mammalian neutrophil elastase, uas tsis pom muaj nyob rau hauv kev tswj peptide thiab lub dehydrating peptide (tshuaj tshem tawm induced lub cev qhuav dej).Qhov kev sim tau rov ua dua peb zaug nrog cov txiaj ntsig zoo sib xws.Piv txwv li, heterologous qhia ntawm ib tug thib ob tshiab \(\bar{d}\)RefSeq = 0.33) pawg ntawm protein biosynthesis elucidates cov kev ua ntawm plaub mature enzymes uas hloov cov 46 amino acid core peptide.Cov seem yog stained raws li qhov chaw ntawm kev hloov kho tau kwv yees los ntawm HR-MS / MS, isotope labeling, thiab NMR tsom xam (Cov Ntaub Ntawv Ntxiv).Dashed xim qhia tau hais tias qhov kev hloov kho tshwm sim ntawm ob qho chaw seem.Daim duab yog muab tso ua ke ntawm ntau heterologous constructs los qhia cov kev ua ntawm tag nrho cov paub tab enzymes nyob rau tib lub nucleus.h, Piv txwv ntawm NMR cov ntaub ntawv rau qaum amide N-methylation.Tag nrho cov txiaj ntsig tau pom hauv daim duab.10 nrog cov ntaub ntawv txuas ntxiv.i, Phylogenetic txoj hauj lwm ntawm cov laus FkbM protein pawg enzyme ntawm tag nrho cov FkbM domains pom nyob rau hauv MIBiG 2.0 database qhia ib enzyme ntawm tsev neeg no nrog N-methyltransferase kev ua (Cov ntaub ntawv ntxiv).Schematic daim duab ntawm BGCs (a, e), precursor peptide lug (b, f), thiab putative chemical structures of natural products (c, g) yog qhia.
Thawj txoj kev RiPP (\(\bar{d}\)MIBiG = 0.41, \(\bar{d}\)RefSeq = 0.29) tsuas yog pom nyob rau hauv hiav txwv tob hom “Ca.E. malaspinii” thiab cov lis dej num rau Peptide-precursor (Fig. 4a, b).Nyob rau hauv no mature enzyme, peb tau txheeb xyuas ib tug functional domain homologous rau lub cev qhuav dej ntawm lantipeptide synthase uas nquag catalyzes phosphorylation thiab tom qab tshem tawm ntawm 43 (Cov ntaub ntawv ntxiv).Yog li ntawd, peb kwv yees tias kev hloov kho ntawm precursor peptide muaj xws li ob-kauj ruam lub cev qhuav dej.Txawm li cas los xij, siv tandem mass spectrometry (MS/MS) thiab nuclear magnetic resonance spectroscopy (NMR), peb tau txheeb xyuas cov polyphosphorylated linear peptide (Fig. 4c).Txawm hais tias kev npaj txhij txog, peb pom ntau cov ntaub ntawv pov thawj los txhawb nws qhov khoom kawg: ob lub tswv yim sib txawv thiab tsis muaj lub cev qhuav dej hauv vitro kev soj ntsuam, kev txheeb xyuas cov seem tseem ceeb hloov pauv hauv catalytic dehydration site ntawm cov enzyme paub tab.tag nrho reconstructed los ntawm "Ca".E. malaspinii genome (cov ntaub ntawv nthuav dav, daim duab 9 thiab cov ntaub ntawv ntxiv) thiab, thaum kawg, kev ua haujlwm lom neeg ntawm cov khoom siv phosphorylated, tab sis tsis yog cov khoom siv tshuaj ua kom lub cev qhuav dej (Fig. 4d).Qhov tseeb, peb pom tias nws nthuav tawm qhov tsis tshua muaj micromolar protease inhibitory kev ua haujlwm tawm tsam neutrophil elastase, piv rau lwm yam khoom siv ntuj tsim nyob rau hauv cov concentration ntau (IC50 = 14.3 μM) 44, txawm hais tias lub luag haujlwm ecological tseem yuav tsum tau piav qhia.Raws li cov txiaj ntsig no, peb xav kom lub npe txoj hauv kev "phospeptin".
Qhov thib ob yog ib txoj hauv kev RiPP txoj kev tshwj xeeb rau 'Ca.Lub genus Eudoremicrobium (\(\bar{d}\)MIBiG = 0.46, \(\bar{d}\)RefSeq = 0.33) tau kwv yees kom encode natural protein cov khoom (Fig. 4e).Txoj hauv kev no yog qhov tshwj xeeb ntawm kev siv tshuab biotechnological vim tias qhov xav tau ceev thiab ntau yam kev hloov pauv tshuaj txawv txawv tsim los ntawm cov enzymes encoded los ntawm luv luv BGCs45.Peb pom tias cov protein no txawv ntawm cov proteins uas tau ua dhau los uas nws tsis muaj ob lub ntsiab NX5N motif ntawm polyceramides thiab lanthionine voj ntawm landornamides 46.Txhawm rau kov yeej cov kev txwv ntawm cov qauv kev nthuav dav dav dav, peb siv lawv nrog rau kev cai Microvirgula aerodenitrificans system los qhia txog plaub txoj hauv kev loj hlob enzymes (txoj kev).Siv kev sib xyaw ua ke ntawm MS/MS, isotope labeling, thiab NMR, peb tau kuaj pom cov enzymes paub tab hauv 46-amino acid core ntawm peptide (Fig. 4f, g, nthuav cov ntaub ntawv, Fig. 10–12 thiab cov ntaub ntawv ntxiv).Ntawm cov enzymes paub tab, peb pom thawj qhov tshwm sim ntawm FkbM O-methyltransferase tsev neeg 47 nyob rau hauv txoj kev RiPP thiab poob nthav pom tias cov enzyme no paub tab qhia cov pob txha N-methylation (Fig. 4h, i thiab cov ntaub ntawv ntxiv).Txawm hais tias qhov kev hloov kho no tau paub hauv cov khoom siv ntuj NRP48, enzymatic N-methylation ntawm amide bonds yog ib qho nyuaj tab sis biotechnologically tseem ceeb cov tshuaj tiv thaiv uas tam sim no tau txaus siab rau RiPP tsev neeg ntawm borosines.Specification 50,51.Kev txheeb xyuas qhov kev ua haujlwm no hauv lwm tsev neeg ntawm cov enzymes thiab RiPP tuaj yeem qhib cov ntawv thov tshiab thiab nthuav dav kev ua haujlwm ntawm cov proteins 52 thiab lawv cov tshuaj muaj ntau haiv neeg.Raws li cov kev hloov kho uas tau txheeb xyuas thiab qhov ntev txawv ntawm cov qauv tsim khoom, peb nthuav tawm txoj hauv kev lub npe "pythonamide".
Qhov kev tshawb pom ntawm ib qho kev npaj txhij txog enzymology hauv ib tsev neeg ua haujlwm ntawm cov enzymes qhia txog kev cog lus ntawm ib puag ncig genomics rau cov kev tshawb pom tshiab, thiab kuj qhia txog qhov txwv tsis pub muaj peev xwm rau kev ua haujlwm raws li kev sib luag homology ib leeg.Yog li, ua ke nrog cov ntawv ceeb toom ntawm non-canonical bioactive polyphosphorylated RiPPs, peb cov txiaj ntsig tau nthuav tawm cov peev txheej-intensive tab sis muaj txiaj ntsig tseem ceeb rau kev siv tshuaj lom neeg biology kom nthuav tawm tag nrho cov kev ua haujlwm nplua nuj, ntau haiv neeg, thiab cov qauv txawv ntawm biochemical compounds.
Ntawm no peb qhia txog ntau yam ntawm biosynthetic muaj peev xwm encoded los ntawm microbes thiab lawv cov ntsiab lus genomic hauv ntiaj teb marine microbiome, ua kom yooj yim rau kev tshawb fawb yav tom ntej los ntawm kev ua kom muaj peev txheej muaj rau cov zej zog tshawb fawb (https://microbiomics.io/ocean/).Peb pom tias ntau ntawm nws cov phylogenetic thiab kev ua haujlwm tshiab tuaj yeem tsuas yog tau los ntawm kev tsim kho MAGs thiab SAGs, tshwj xeeb tshaj yog nyob rau hauv cov zej zog tsis zoo uas tuaj yeem coj mus rau yav tom ntej bioprospecting.Txawm hais tias peb yuav tsom mus rau ntawm 'Ca.Eudormicrobiaceae" raws li ib tug kab mob tshwj xeeb tshaj yog biosynthetically "tshaj", ntau ntawm BGCs kwv yees nyob rau hauv undiscovered microbiota yuav encode yav dhau los undescribed enzymologies uas yields tebchaw nrog ib puag ncig thiab / los yog biotechnologically ua tseem ceeb.
Metagenomic datasets los ntawm cov dej hiav txwv loj thiab lub sij hawm series kev tshawb fawb nrog txaus sequencing qhov tob tau muab tso rau kom loj tshaj kev pab cuam ntawm lub ntiaj teb no marine microbial zej zog nyob rau hauv dej hiav txwv basins, sib sib zog nqus txheej thiab dhau sij hawm.Cov ntaub ntawv no (Cov Lus Qhia Ntxiv 1 thiab Daim duab 1) suav nrog cov metagenomics los ntawm cov qauv sau hauv hiav txwv ntawm Tara (viral enriched, n = 190; prokaryotic enriched, n = 180) 12,22 thiab BioGEOTRACES expedition (n = 480).Hawaiian Oceanic Time Series (HOT, n = 68), Bermuda-Atlantic Time Series (BATS, n = 62)21 thiab Malaspina Expedition (n = 58)23.Sequencing nyeem los ntawm tag nrho cov metagenomic fragments tau lim kom zoo siv BBMap (v.38.71) los ntawm kev tshem tawm sequencing adapters los ntawm kev nyeem, tshem tawm nyeem mapped rau zoo tswj sequences (PhiX genomes), thiab siv trimq = 14, maq = 20 pov tseg tsis zoo nyeem ntawv, maxns = 0 thiab minlength = 45. Cov kev soj ntsuam tom qab tau khiav los yog sib xyaw nrog QC nyeem yog tias tau teev tseg (bbmerge.sh minoverlap=16).QC nyeem tau zoo li qub (bbnorm.sh lub hom phiaj = 40, minddepth = 0) ua ntej tsim siv metaSPAdes (v.3.11.1 lossis v.3.12 yog xav tau)53.Qhov tshwm sim scaffold contigs (tom qab no hu ua scaffolds) thaum kawg tau lim los ntawm qhov ntev (≥1 kb).
Cov 1038 cov qauv ntsuas tau muab faib ua pawg, thiab rau txhua pawg ntawm cov qauv, kev tswj hwm kev tswj hwm zoo ntawm txhua tus qauv raug muab sib xyaw rau cov kab ke ntawm txhua tus qauv sib cais, ua rau muaj cov nram qab no ntawm pawg sib txuas sib txuas nyeem: Tara Marine Viruses - Enriched (190 × 190), Prokaryotes Enriched (180 × 180), BioGEOTRACES, HOT thiab BATS (610 × 610) thiab Malaspina (58 × 58).Daim ntawv qhia tau ua tiav siv Burrows-Wheeler-Aligner (BWA) (v.0.7.17-r1188)54 uas tso cai rau kev nyeem ntawv kom phim rau qhov chaw thib ob (siv tus chij -a).Alignments raug lim kom tsawg kawg yog 45 lub hauv paus ntev, muaj ≥97% tus kheej, thiab ncua ≥80% nyeem.Cov ntaub ntawv BAM uas tau tshwm sim tau ua tiav siv cov ntawv jgi_summarize_bam_contig_depths rau MetaBAT2 (v.2.12.1)55 los muab kev pab cuam sib piv rau txhua pawg.Thaum kawg, cov brackets tau muab faib ua pawg kom muaj kev nkag siab zoo los ntawm tus kheej khiav MetaBAT2 ntawm txhua qhov qauv nrog -minContig 2000 thiab -maxEdges 500. Peb siv MetaBAT2 tsis yog ib qho kev sib tw boxer vim nws tau pom nyob rau hauv kev xeem ywj pheej los ua qhov zoo tshaj plaws hauv ib lub boxer.thiab 10 mus rau 50 npaug sai dua li lwm cov boxers uas nquag siv 57.Txhawm rau ntsuam xyuas cov txiaj ntsig ntawm kev sib raug zoo ntawm kev sib raug zoo, ib qho kev xaiv los ntawm cov qauv ntawm metagenomics (10 rau txhua qhov ntawm ob daim ntawv teev dej hiav txwv Tara, 10 rau BioGEOTRACES, 5 rau txhua lub sijhawm, thiab 5 rau Malaspina) ntxiv rau cov qauv siv xwb.Cov qauv sab hauv raug muab ua pab pawg kom tau txais cov ntaub ntawv pov thawj.(Cov ntaub ntawv ntxiv).
Ntxiv (sab nrauv) genomes tau suav nrog hauv kev tshuaj ntsuam tom ntej, uas yog 830 manually xaiv MAGs los ntawm ib pawg ntawm Tara Oceans26 dataset, 5287 SAGs los ntawm GORG20 dataset, thiab cov ntaub ntawv los ntawm MAR database (MarDB v. 4) los ntawm 1707 cais REFs thiab 682 SAGs) 27. Rau MarDB dataset, genomes raug xaiv raws li muaj metadata yog tias hom qauv sib phim cov lus qhia hauv qab no: '[S|s]ingle.?[C|c]ell|[C|c]ulture| [I|i] cais'.
Qhov zoo ntawm txhua lub ntim ntim thiab cov genomes sab nraud tau soj ntsuam siv CheckM (v.1.0.13) thiab Anvi'o's Lineage Workflow (v.5.5.0) 58,59.Yog tias CheckM lossis Anvi'o tshaj tawm ≥50% ua tiav / ua tiav thiab ≤10% kev sib kis / rov ua dua, ces txuag cov hlwb metagenomic thiab cov genomes sab nraud rau kev tshuaj xyuas tom qab.Cov qhab nias no tau muab tso ua ke rau hauv qhov kev ua tiav (mcpl) thiab txhais tau tias muaj kev sib kis (mctn) kom faib cov genome zoo raws li cov txheej txheem hauv zej zog 60 raws li hauv qab no: siab zoo: mcpl ≥ 90% thiab mctn ≤ 5%;zoo zoo: mcpl ≥ 70%, mctn ≤ 10%, nruab nrab zoo: mcpl ≥ 50% thiab mctn ≤ 10%, ncaj ncees zoo: mcpl ≤ 90% lossis mctn ≥ 10%.Cov genomes lim tau ces sib txheeb nrog cov qhab nia zoo (Q thiab Q') raws li hauv qab no: Q = mcpl – 5 x mctn Q' = mcpl – 5 x mctn + mctn x (strain variability)/100 + 0.5 x log[N50] .(ua tiav hauv dRep61).
Txhawm rau tso cai kev sib piv ntawm cov ntaub ntawv sib txawv thiab hom genome (MAG, SAG thiab REF), 34,799 genomes tau dereferenced raws li genome-wide nruab nrab nucleotide tus kheej (ANI) siv dRep (v.2.5.4).Rov ua dua)61 ​​nrog 95% ANI thresholds28,62 (-comp 0 -con 1000 -sa 0.95 -nc 0.2) thiab ib daim ntawv theej cim cov noob siv SpecI63 muab genome clustering ntawm qib hom.Ib tus neeg sawv cev genome raug xaiv rau txhua pawg dRep raws li qhov qhab nia siab tshaj plaws (Q') tau hais tseg saum toj no, uas tau suav tias yog tus sawv cev ntawm hom.
Txhawm rau ntsuas qhov ntsuas nrawm, BWA (v.0.7.17-r1188, -a) tau siv los qhia tag nrho 1038 pawg ntawm metagenomic nyeem nrog 34,799 genomes muaj nyob hauv OMD.Kev tswj xyuas kev nyeem tau zoo tau kos npe nyob rau hauv ib qho kawg hom thiab cov txiaj ntsig kev sib tw tau raug lim kom khaws cia tsuas yog kev sib tw ≥45 bp ntev.thiab tus kheej ≥ 95%.Cov zaub piv rau txhua tus qauv yog qhov feem pua ​​​​ntawm kev nyeem ntawv ntxiv tom qab pom tau muab faib los ntawm tag nrho cov kev tswj xyuas kev nyeem ntawv zoo.Siv tib txoj hauv kev, txhua qhov ntawm 1038 metagenomes raug txo mus rau 5 lab qhov ntxig (cov ntaub ntawv nthuav dav, daim duab 1c) thiab sib phim rau GORG SAG hauv OMD thiab hauv txhua GEM16.Tus nqi ntawm MAGs rov qab los ntawm cov dej hiav txwv hauv GEM16 catalog tau txiav txim siab los ntawm cov lus nug tseem ceeb ntawm qhov chaw metagenomic, xaiv cov qauv dej hiav txwv (xws li, tsis zoo li marine sediments).Tshwj xeeb, peb xaiv "dej" ua "ecosystem_category", "marine" li "ecosystem_type", thiab lim "habitat" li "dej hiav txwv", "marine", "marine dej hiav txwv", "pelagic marine", "marine water", "Hiav txwv", "Hiav txwv dej", "Ntog hiav txwv dej", "Ntog hiav txwv dej".Qhov no ua rau 5903 MAGs (734 zoo) faib tshaj 1823 OTUs (saib ntawm no).
Prokaryotic genomes tau sau tseg los ntawm taxonomically siv GTDB-Tk (v.1.0.2)64 nrog lub neej ntawd tsis muaj hom phiaj GTDB r89 version 13. Anvi'o tau siv los txheeb xyuas cov genomes eukaryotic raws li kev kwv yees sau npe thiab nco qab ≥50% thiab redundancy ≤ 10%.Lub taxonomic annotation ntawm ib hom yog txhais raws li ib tug ntawm nws cov neeg sawv cev genomes.Nrog rau kev zam ntawm eukaryotes (148 MAG), txhua lub genome yog thawj zaug ua haujlwm nrog cov lus piav qhia siv prokka (v.1.14.5)65, naming tiav cov noob, txhais "archaea" lossis "cov kab mob" tsis raws li xav tau, uas tseem tau tshaj tawm rau cov tsis- coding noob.thiab CRISPR cheeb tsam, ntawm lwm yam genomic nta.Tshaj tawm cov noob kwv yees los ntawm kev txheeb xyuas universal single-copy marker genes (uscMG) siv fetchMG (v.1.2)66, muab pawg ortholog thiab nug siv emapper (v.2.0.1)67 raws li eggNOG (v.5.0)68.KEGG database (tshaj tawm Lub Ob Hlis 10, 2020) 69. Cov kauj ruam kawg tau ua los ntawm kev sib piv cov proteins rau KEGG database siv DIAMOND (v.0.9.30)70 nrog cov lus nug thiab cov ntsiab lus ntawm ≥70%.Cov txiaj ntsig tau raug lim ntxiv raws li NCBI Prokaryotic Genome Annotation Pipeline71 raws li bitrate ≥ 50% ntawm qhov siab tshaj qhov xav tau bitrate (txuas nws tus kheej).Gene sequences kuj tau siv los ua cov tswv yim los txheeb xyuas BGCs hauv cov genome siv antiSMASH (v.5.1.0)72 nrog rau qhov tsis zoo thiab qhov sib txawv ntawm pawg tawg.Tag nrho cov genomes thiab cov lus piav qhia tau muab tso ua ke rau hauv OMD nrog rau cov ntsiab lus metadata muaj nyob hauv lub vev xaib (https://microbiomics.io/ocean/).
Zoo ib yam li tau piav qhia yav dhau los 12,22 peb siv CD-HIT (v.4.8.1) rau pawg> 56.6 lab protein-coding noob los ntawm cov kab mob thiab cov archaeal genomes los ntawm OMD rau 95% yog tus kheej thiab cov noob luv (90% kev pab them nqi) 73 txog > 17.7 lab gene pawg.Cov kab ke ntev tshaj plaws tau raug xaiv los ua tus sawv cev gene rau txhua pawg gene.Lub 1038 metagenomes tau muab sib xyaw rau > 17.7 lab BWA (-a) pawg tswv cuab thiab cov ntaub ntawv BAM tau raug lim kom khaws cia tsuas yog kev sib raug zoo nrog ≥95% feem pua ​​​​ntawm tus kheej thiab ≥45 qhov kev sib dhos.Length-normalized gene abundance yog xam los ntawm thawj suav inserts los ntawm qhov zoo tshaj plaws kev sib raug zoo tshwj xeeb thiab tom qab ntawd, rau fuzzy-mapped inserts, ntxiv fractional suav rau cov hom phiaj sib txuas rau lawv cov naj npawb ntawm cov cim tshwj xeeb.
Cov genomes los ntawm OMD nthuav dav (nrog MAGs ntxiv los ntawm "Ca. Eudormicrobiaceae", saib hauv qab no) tau ntxiv rau mOTUs74 metagenomic tsom xam cov cuab yeej database (v.2.5.1) los tsim cov ntaub ntawv txuas ntxiv mOTU siv.Tsuas yog rau ib qho genomes (23,528 genomes) tau dim ntawm kaum uscMGs.Kev nthuav dav ntawm cov ntaub ntawv tau ua rau 4,494 pawg ntxiv ntawm qib hom.1038 metagenomes tau txheeb xyuas siv qhov tsis zoo mOTU (v.2).Tag nrho ntawm 989 genomes muaj nyob rau hauv 644 mOTU pawg (95% REF, 5% SAG thiab 99.9% belongs rau MarDB) tsis pom los ntawm mOTU profile.Qhov no qhia txog ntau qhov chaw ntawm kev sib cais marine ntawm MarDB genomes (feem ntau ntawm cov genomes uas tsis tau kuaj pom muaj feem cuam tshuam nrog cov kab mob sib cais los ntawm sediments, marine hosts, thiab lwm yam).Txhawm rau txuas ntxiv tsom mus rau qhov qhib dej hiav txwv ib puag ncig hauv txoj kev tshawb no, peb tshem tawm lawv los ntawm kev txheeb xyuas qis qis tshwj tsis yog lawv tau kuaj pom lossis suav nrog hauv cov ntaub ntawv txuas ntxiv mOTU tsim hauv txoj kev tshawb no.
Tag nrho BGCs los ntawm MAG, SAG thiab REF hauv OMD (saib saum toj no) tau ua ke nrog BGCs pom nyob rau hauv tag nrho cov metagenomic scaffolds (antiSMASH v.5.0, default parameters) thiab characterized siv BiG-SLICE (v.1.1) (PFAM domain )75.Raws li cov yam ntxwv no, peb suav tag nrho cov cosine nrug ntawm BGCs thiab muab faib ua pawg (txhais tau tias txuas) rau hauv GCF thiab GCC siv qhov kev ncua deb ntawm 0.2 thiab 0.8 raws.Cov theem pib no yog kev hloov pauv ntawm qhov pib siv yav dhau los siv Euclidean nrug75 ua ke nrog cosine nrug, uas txo qee qhov yuam kev hauv thawj BiG-SLICE pawg tswv yim (Cov Ntaub Ntawv Ntxiv).
BGCs tau raug lim kom khaws cia tsuas yog ≥5 kb encoded ntawm scaffolds los txo qhov kev pheej hmoo ntawm fragmentation raws li yav dhau los tau piav qhia 16 thiab tshem tawm MarDB REFs thiab SAGs tsis pom hauv 1038 metagenomes (saib saum toj).Qhov no tau ua rau tag nrho ntawm 39,055 BGCs raug encoded los ntawm OMD genome, nrog rau ntxiv 14,106 txheeb xyuas ntawm cov kab mob metagenomic (piv txwv li tsis ua ke rau hauv MAGs).Cov "metagenomic" BGCs tau siv los kwv yees qhov feem pua ​​​​ntawm marine microbiome biosynthesis muaj peev xwm tsis ntes hauv cov ntaub ntawv (Cov Ntaub Ntawv Ntxiv).Txhua BGC tau ua haujlwm zoo raws li kev kwv yees cov khoom lag luam uas tau hais los ntawm cov khoom lag luam los tiv thaiv SMASH lossis coarser pawg tau teev tseg hauv BiG-SCAPE76.Txhawm rau tiv thaiv kev tsis sib haum xeeb hauv kev ua kom muaj nuj nqis (taxonomic thiab kev ua haujlwm ntawm GCC / GCF, qhov deb ntawm GCF thiab GCC rau kev siv cov ntaub ntawv, thiab metagenomic abundance ntawm GCF), los ntawm kev khaws cia tsuas yog qhov ntev tshaj plaws BGC ib GCF rau txhua hom, 39,055 BGCs tau ntxiv deduplicated, ua rau tag nrho ntawm 17,689 BGC.
Qhov tshiab ntawm GCC thiab GCF tau soj ntsuam raws li qhov kev ncua deb ntawm cov ntaub ntawv suav (RefSeq database hauv BiG-FAM) 29 thiab qhov kev sim ua pov thawj (MIBIG 2.0) 30 BGC.Rau txhua tus ntawm 17,689 tus neeg sawv cev BGCs, peb xaiv qhov tsawg tshaj plaws cosine nrug rau cov ntaub ntawv sib tham.Qhov kev ncua deb tsawg kawg no yog qhov nruab nrab (txhais tau tias) raws li GCF lossis GCC, raws li qhov tsim nyog.GCF raug suav hais tias yog qhov tshiab yog qhov kev ncua deb ntawm cov ntaub ntawv ntau dua 0.2, uas sib haum rau kev sib cais zoo tshaj plaws ntawm (nruab nrab) GCF thiab cov ntaub ntawv siv.Rau GCC, peb xaiv 0.4, uas yog ob zaug ntawm qhov pib uas tau teev tseg los ntawm GCF, txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm rau txhawm
Lub metagenomic abundance ntawm BGC yog kwv yees li qhov nruab nrab abundance ntawm nws biosynthetic genes (raws li txiav txim los ntawm anti-SMASH) muaj los ntawm cov noob theem profiles.Lub metagenomic abundance ntawm txhua GCF los yog GCC yog ces xam raws li tus lej ntawm cov neeg sawv cev BGCs (tawm ntawm 17,689).Cov ntaub ntawv qhia kev nplua nuj no tau ua raws li ib txwm ua rau cov cellular muaj pes tsawg leeg siv ib tus qauv mOTU suav, uas tseem suav nrog kev siv zog ua ntu zus (cov ntaub ntawv nthuav dav, daim duab 1d).Qhov feem ntau ntawm GCF lossis GCC raug xam raws li feem pua ​​​​ntawm cov qauv uas muaj ntau> 0.
Qhov kev ncua deb ntawm Euclidean ntawm cov qauv raug xam los ntawm qhov normalized GCF profile.Cov kev ncua deb no tau txo qhov loj me me siv UMAP77 thiab qhov tshwm sim embeddings tau siv rau qhov tsis muaj kev saib xyuas raws li kev sib koom ua ke siv HDBSCAN78.Qhov zoo tshaj plaws tus naj npawb ntawm cov ntsiab lus rau ib pawg (thiab yog li tus naj npawb ntawm pawg) siv los ntawm HDBSCAN yog txiav txim siab los ntawm kev ua kom muaj qhov ua tau zoo ntawm pawg tswv cuab.Cov pawg uas tau txheeb xyuas (thiab ib qho kev sib npaug sib npaug ntawm cov pawg no los suav rau kev tsis ncaj ncees hauv kev txheeb xyuas qhov sib txawv ntawm qhov sib txawv (PERMANOVA)) tau sim rau qhov tseem ceeb tiv thaiv qhov tsis tau txo qis Euclidean nrug siv PERMANOVA.Qhov nruab nrab genome loj ntawm cov qauv raug xam raws li qhov txheeb ze ntawm mOTU thiab kwv yees qhov loj ntawm cov genomes ntawm cov tswv cuab.Tshwj xeeb, qhov nruab nrab genome loj ntawm txhua mOTU tau kwv yees li qhov nruab nrab ntawm cov genome loj ntawm nws cov tswv cuab kho kom tiav (tom qab lim) (piv txwv li, 75% ua tiav genome nrog qhov ntev ntawm 3 Mb muaj qhov hloov kho loj ntawm 4. Mb).rau nruab nrab genomes nrog kev ncaj ncees ≥70%.Qhov nruab nrab genome loj rau txhua tus qauv raug xam raws li qhov sib npaug ntawm mOTU genome qhov ntau thiab tsawg uas hnyav los ntawm cov txheeb ze ntau.
Ib txheej lim dej ntawm genome-encoded BGCs hauv OMD yog pom nyob rau hauv cov kab mob thiab archaeal GTDB cov ntoo (hauv ≥5 kb lub moj khaum, tsis suav nrog REF thiab SAG MarDB tsis pom nyob rau hauv 1038 metagenomes, saib saum toj no) thiab lawv cov khoom kwv yees raws li cov phylogenetic txoj hauj lwm ntawm genome (saib saum toj).Peb thawj zaug txo cov ntaub ntawv los ntawm hom, siv cov genome nrog cov BGCs feem ntau hauv cov tsiaj ntawd ua tus sawv cev.Rau kev pom, cov neeg sawv cev tau muab faib ua pawg ntoo, thiab dua, rau txhua lub celled clade, cov genome uas muaj cov BGC ntau tshaj plaws tau raug xaiv los ua tus sawv cev.BGC-enriched hom (tsawg kawg yog ib lub genome nrog > 15 BGCs) tau soj ntsuam ntxiv los ntawm kev suav cov Shannon Diversity Index rau cov khoom hom encoded hauv cov BGCs.Yog tias txhua yam khoom lag luam kwv yees zoo ib yam, cov tshuaj hybrids thiab lwm yam BGCs complex (raws li kev kwv yees los ntawm anti-SMAH) raug suav hais tias yog cov khoom lag luam tib yam, tsis hais lawv qhov kev txiav txim hauv pawg (xws li protein-bacteriocin thiab bacteriocin-proteoprotein fusion . lub cev).hybrid).
Tseem tshuav DNA (kwv yees li 6 ng) los ntawm Malaspina qauv MP1648, sib xws rau cov qauv lom neeg SAMN05421555 thiab sib phim rau Illumina SRR3962772 metagenomic nyeem teeb rau luv luv nyeem, ua tiav raws li PacBio sequencing raws tu qauv nrog ultra-tsawg ampliobell cov khoom siv PacNA Cov khoom siv (100-980-000) thiab SMRTbell Express 2.0 template npaj khoom siv (100-938-900).Luv luv, qhov seem DNA raug txiav, kho thiab ua kom huv (ProNex hlaws) siv Covaris (g-TUBE, 52104).Purified DNA yog tom qab ntawd raug rau cov tsev qiv ntawv npaj, nthuav tawm, ua kom huv (ProNex hlaws) thiab xaiv qhov loj me (> 6 kb, Blue Pippin) ua ntej cov kauj ruam kawg purification (ProNex hlaws dai) thiab sib sau ua ke ntawm Sequel II platform.
Reconstruction ntawm thawj ob ca.Rau MAG Eremiobacterota, peb tau txheeb xyuas 6 qhov ANIs ntxiv > 99% (cov no suav nrog hauv daim duab 3), uas tau pib lim raws li cov qhab nia sib kis (tom qab tau txheeb xyuas cov noob duplications, saib hauv qab).Peb kuj pom ib lub tais sau tias "Ca".Eremiobacterota" los ntawm ntau yam kev tshawb fawb23 thiab siv lawv ua ke nrog yim MAGs los ntawm peb txoj kev tshawb fawb raws li kev siv rau metagenomic nyeem los ntawm 633 eukaryotic enriched (> 0.8 µm) cov qauv siv BWA (v.0.7.17) Ref -r1188, - tus chij) rau downsampled daim ntawv qhia (5 lab nyeem).Raws li kev nthuav qhia tshwj xeeb hauv daim duab qhia chaw ( lim los ntawm 95% kev sib koom ua ke ntawm tus kheej thiab 80% nyeem cov kev pab), 10 metagenomes (xav tau kev pab them nqi ≥5 ×) raug xaiv los ua ke thiab ntxiv 49 metagenomes (xav tau kev pab them nqi ≥1 ×) rau cov ntsiab lus sib txheeb.Siv tib yam tsis xws li saum toj no, cov qauv no tau ntim thiab 10 ntxiv 'Ca's tau ntxiv.MAG Eremiobacterota tau rov qab los.Cov 16 MAGs no (tsis suav ob qho uas twb muaj lawm hauv cov ntaub ntawv) coj tag nrho cov genomes hauv OMD nthuav dav rau 34,815.MAGs tau muab cov qeb taxonomic raws li lawv qhov kev sib raug zoo thiab txoj haujlwm hauv GTDB.18 MAGs tau dereplicated siv dRep rau hauv 5 hom (intraspecific ANI> 99%) thiab 3 genera (intrageneric ANI 85% mus rau 94%) nyob rau hauv tib tsev neeg79.Cov neeg sawv cev ntawm cov tsiaj tau raug xaiv los ntawm kev ncaj ncees, kev sib kis, thiab N50.Cov lus pom zoo nomenclature muaj nyob rau hauv Cov Lus Qhia Ntxiv.
Ntsuam xyuas kev ncaj ncees thiab kev sib kis ntawm 'Ca.MAG Eremiobacterota, peb tau soj ntsuam lub xub ntiag ntawm uscMG, nrog rau cov caj ces- thiab cov npe tshwj xeeb ib qho cim cim npe siv los ntawm CheckM thiab Anvi'o.Qhov kev txheeb xyuas ntawm 2 qhov sib npaug ntawm 40 uscMGs tau lees paub los ntawm phylogenetic reconstruction (saib hauv qab) los txiav txim tawm tej yam kev kis kab mob (qhov no sib raug rau 5% raws li cov 40 cov cim npe no).Ib qho kev kawm ntxiv ntawm tsib tus neeg sawv cev MAGs 'Ca.Qib qis ntawm cov kab mob hauv cov genomes uas tau tsim dua tshiab no tau lees paub rau Eremiobacterota hom siv kev sib tham sib Anvi'o interface raws li kev nplua nuj thiab kev sib txuas ua ke (Cov Lus Qhia Ntxiv)59.
Rau kev tsom xam phylogenomic, peb xaiv tsib tus neeg sawv cev MAGs "Ca".Eudormicrobiaceae, txhua hom "Ca.Lub genome ntawm Eremiobacterota thiab cov tswv cuab ntawm lwm cov phyla (xws li UBP13, Armatimonadota, Patescibacteria, Dormibacterota, Chloroflexota, Cyanobacteria, Actinobacteria thiab Planctomycetota) muaj los ntawm GTDB (r89)13.Tag nrho cov genomes no tau sau tseg raws li tau piav qhia yav dhau los rau ib daim ntawv luam cim noob rho tawm thiab BGC annotation.GTDB genomes tau txais kev txuag raws li kev ncaj ncees thiab kev sib kis saum toj no.Kev tshuaj xyuas Phylogenetic tau ua tiav siv Anvi'o Phylogenetics59 workflow.Cov ntoo tau tsim los siv IQTREE (v.2.0.3) (kev xaiv ua ntej thiab -bb 1000)80 ntawm kev sib dhos ntawm 39 tandem ribosomal proteins txheeb xyuas los ntawm Anvi'o (MUSCLE, v.3.8.1551)81.Nws txoj haujlwm raug txo.kom npog tsawg kawg 50% ntawm genome82 thiab Planctomycecota tau siv los ua ib pawg neeg tawm raws li GTDB ntoo topology.Ib tsob ntoo ntawm 40 uscMGs tau tsim los siv tib cov cuab yeej thiab cov kev txwv.
Peb siv Traitar (v.1.1.2) nrog lub neej ntawd tsis (phenotype, los ntawm nucleotides)83 los kwv yees cov kab mob microbial.Peb tshawb nrhiav qhov muaj peev xwm ua neej nyob raws li kev tsim ua ntej predatory index84 uas nyob ntawm cov ntsiab lus ntawm cov protein-coding noob hauv genome.Tshwj xeeb, peb siv DIAMOND los sib piv cov proteins hauv cov genome tawm tsam OrthoMCL database (v.4) 85 siv cov kev xaiv -ntau-sensive -id 25 -query-cover 70 -subject-cover 70 -top 20 THIAB suav cov noob sib raug rau cov cim genes rau predators thiab tsis-predators.Qhov ntsuas yog qhov sib txawv ntawm tus naj npawb ntawm predatory thiab tsis-predatory cim.Raws li kev tswj hwm ntxiv, peb kuj tau txheeb xyuas "Ca" genome.Entotheonella TSY118 yog raws li nws cov koom haum nrog Ca.Eudoremicrobium (loj genome loj thiab muaj peev xwm biosynthetic).Tom ntej no, peb tau sim cov kev sib txuas ntawm cov tsiaj nyeg thiab cov tsiaj txhu uas tsis yog neeg tua tsiaj thiab cov peev xwm biosynthetic ntawm Ca.Eudormicrobiaceae" thiab pom tias tsis muaj ntau tshaj li ib lub noob (los ntawm txhua hom cim cim, piv txwv li cov tsiaj txhu / tsis yog tsiaj txhu) sib tshooj nrog BGC, qhia tias BGC tsis cuam tshuam cov cim kev ua phem.Ntxiv cov lus piav qhia genomic ntawm scrambled replicaons tau ua tiav siv TXSSCAN (v.1.0.2) txhawm rau tshuaj xyuas cov txheej txheem zais cia, pili, thiab flagella86.
Tsib tus neeg sawv cev 'Ca's tau ua los ntawm daim ntawv qhia 623 metatranscriptomes los ntawm prokaryotic thiab eukaryotic enrichment feem ntawm Tara oceans22,40,87 (siv BWA, v.0.7.17-r1188, -a chij).Eudormicrobiaceae genome.BAM cov ntaub ntawv tau ua tiav nrog FeatureCounts (v.2.0.1)88 tom qab 80% nyeem cov kev pab them nqi thiab 95% kev txheeb xyuas tus kheej (nrog cov kev xaiv featureCounts -primary -O -fraction -t CDS,tRNA -F GTF -g ID -p) suav cov tus naj npawb ntawm inserts ib gene.Cov duab qhia chaw tsim tau zoo li qub rau cov noob ntev thiab cov cim gene abundance mOTU (ntev-normalized nruab nrab qhov nruab nrab suav rau cov noob nrog suav suav> 0) thiab log-hloov mus rau 22.74 kom tau txais cov txheeb ze qhia ib lub xovtooj ntawm txhua qib gene, uas tseem piav qhia variability ntawm tus qauv mus rau tus qauv thaum lub sij hawm sequencing.Xws li qhov piv txwv tso cai rau kev sib piv, txo cov teeb meem sib xyaw thaum siv cov ntaub ntawv txheeb ze.Tsuas yog cov qauv nrog > 5 ntawm 10 mOTU cov cim cim noob tau raug txiav txim siab rau kev tshuaj xyuas ntxiv kom tso cai rau qhov loj txaus ntawm cov genome los kuaj pom.
Lub normalized transcriptome profile ntawm 'Ca.E. taraoceanii tau raug txo kom muaj nuj nqis siv UMAP thiab cov txiaj ntsig sawv cev tau siv rau kev tsis saib xyuas pawg siv HDBSCAN (saib saum toj no) los txiav txim cov xwm txheej qhia.PERMANOVA ntsuas qhov tseem ceeb ntawm qhov sib txawv ntawm cov pawg uas tau txheeb xyuas hauv qhov qub (tsis txo qis) qhov chaw nyob deb.Kev qhia txawv ntawm cov xwm txheej no tau sim hla cov genome (saib saum toj no) thiab 201 KEGG txoj hauv kev tau txheeb xyuas hauv 6 pawg ua haujlwm, xws li: BGC, secretion system thiab flagellar genes los ntawm TXSSCAN, degradation enzymes (protease thiab peptidases), thiab predatory thiab non- predatory noob.predatory index markers.Rau txhua tus qauv, peb xam qhov nruab nrab ntawm cov lus qhia rau txhua chav kawm (nco ntsoov tias BGC qhia nws tus kheej yog xam raws li qhov nruab nrab ntawm biosynthetic genes rau BGC) thiab kuaj qhov tseem ceeb thoob plaws xeev (Kruskal-Wallis kuaj kho rau FDR).
Cov noob hluavtaws tau yuav los ntawm GenScript thiab PCR primers tau yuav los ntawm Microsynth.Phusion polymerase los ntawm Thermo Fisher Scientific tau siv rau DNA amplification.NucleoSpin plasmids, NucleoSpin gel thiab PCR purification cov khoom siv los ntawm Macherey-Nagel tau siv rau DNA purification.Kev txwv enzymes thiab T4 DNA ligase tau yuav los ntawm New England Biolabs.Cov tshuaj uas tsis yog isopropyl-β-d-1-thiogalactopyranoside (IPTG) (Biosynth) thiab 1,4-dithiothreitol (DTT, AppliChem) tau yuav los ntawm Sigma-Aldrich thiab siv yam tsis muaj kev lim dej ntxiv.Cov tshuaj tua kab mob chloramphenicol (Cm), spectinomycin dihydrochloride (Sm), ampicillin (Amp), gentamicin (Gt), thiab carbenicillin (Cbn) tau yuav los ntawm AppliChem.Bacto Tryptone thiab Bacto Poov xab Extract media Cheebtsam tau yuav los ntawm BD Biosciences.Trypsin rau sequencing yog yuav los ntawm Promega.
Gene sequences tau muab rho tawm los ntawm anti-SMASH kwv yees BGC 75.1.E. malaspinii (Cov ntaub ntawv ntxiv).
Cov noob embA (locus, MALA_SAMN05422137_METAG-framework_127-gene_5), embM (locus, MALA_SAMN05422137_METAG-framework_127-gene_4), thiab embAM (suav nrog thaj tsam ntawm kev sib txuas nrog UC7) uas tsis muaj kev sib txuas ntawm cov kab ke (piv txwv li UC) codons optimized rau kev qhia hauv E thaum.Cov noob embA tau raug subcloned rau hauv thawj ntau qhov chaw cloning (MCS1) ntawm pACYCDuet-1(CmR) thiab pCDFDuet-1(SmR) nrog BamHI thiab HindIII cleavage sites.Cov noob embM thiab embMopt (codon-optimized) tau subcloned rau hauv MCS1 pCDFDuet-1(SmR) nrog BamHI thiab HindIII thiab muab tso rau hauv qhov thib ob ntau qhov chaw cloning ntawm pCDFDuet-1(SmR) thiab pRSFDuet-1(KanR) (MCS2) nrog NdeI/ChoI.Lub embAM cassette tau subcloned rau hauv pCDFDuet1 (SmR) nrog BamHI thiab HindIII cleavage sites.Lub orf3/embI noob (locus, MALA_SAMN05422137_METAG-scaffold_127-gene_3) tau tsim los ntawm kev sib tshooj txuas PCR siv cov primers EmbI_OE_F_NdeI thiab EmbI_OE_R_XhoI, zom nrog NdeI / ated rau pMCet1 re-MCD (ligested) kev txwv enzymes (Ntxiv rooj).6).Kev txwv enzyme digestion thiab ligation tau ua raws li cov chaw tsim khoom raws tu qauv (New England Biolabs).