I-DNA / i-RNA yokulandelelana -I-PacBio Sequencer
Iimbonakalo
Iindlela ezimbini zokulandelelana kwi-sequencer ye-PacBio: Ukufunda okude okuqhubekayo (CLR) kunye neSetyhula Consensus Read (CCS)
| Imo Yolandelelwano | Ubungakanani beThala leeNcwadi | Idatha yethiyoriYield (NgeSeli nganye) | Isiseko esinyeUkuchaneka | Usetyenziso |
| I-CLR | 20Kb, 30Kb, njl. | I-80 Gb ukuya kwi-130 Gb | Malunga.85% | De novo, ukufowuna kwe-SV, njl. |
| CCS | 15-20 KB | 14 ukuya 40 Gb/Cell (Sequel II) 70 ukuya 110 Gb/Cell (Revio) Kuxhomekeke kwiisampuli | Malunga.99% | De novo, SNP/Indel/SV ukufowuna, Iso-Seq, |
Ukuthelekisa ukusebenza kunye neempawu zeRevio kunye neSequel II
| Imigaqo | Inkqubo ye-Sequel II | Inkqubo yeRevio | Nyusa |
| Ukuxinana okuphezulu | Zizigidi ezisi-8 zee-ZMWs | 25 yezigidi ze-ZMWs | 3x |
| Izigaba ezizimeleyo | 1 | 4 | 4x |
| Amaxesha amafutshane okubaleka | iiyure ezingama-30 | iiyure ezingama-24 | 1.25x |
| 30X HiFi iigenomes zabantu / ngonyaka | 88 | 1,300 | 15x ngokupheleleyo |
Iinkonzo eziluncedo
● Ngaphezulu kweminyaka eyi-8 yamava kwi-PacBio yokulandelelana kweqonga kunye namawaka eeprojekthi ezivaliweyo ezineentlobo ezahlukeneyo.
● Ixhotyiswe ngokupheleleyo kunye neeplatifti zePacBio Sequencing zamva nje, iRevio yokuqinisekisa ulandelelwano olwaneleyo lokuphuma.
● Ixesha elikhawulezayo lokujika, isivuno esiphezulu sedatha kunye nedatha echaneke ngakumbi.
● Igalelo kumakhulu eempembelelo eziphezulu zePacBio ezipapashiweyo.
IiMfuno zeSampulu
| Uhlobo lweSampuli | Isixa | Ugxininiso (Qubit®) | Umthamo | Ubunyulu | Abanye |
| Genomic DNA | Kuxhomekeke kwiMfuno yeDatha | ≥50 ng/μl | ≥15μl | OD260/280=1.7-2.2; OD260/230=1.8-2.5; Incopho ecacileyo kwi-260nm,akukho ungcoliseko | Ugxininiso kufuneka lulinganiswe ngeQubit kunye neQubit / Nanopore = 0.8-2.5 |
| Iyonke i-RNA | ≥1.2μg | ≥120 ng/μl | ≥15μl | OD260/280=1.7-2.5; OD260/230=0.5-2.5;akukho ungcoliseko | Ixabiso le-RIN ≥7.5 5≥28S/18S≥1 |
Ukuhanjiswa kwenkonzo
Ukulungiselela isampuli
Ukwakhiwa kwethala leencwadi
Ukulandelelanisa
Uhlalutyo lwedatha
Unikezelo lweprojekthi
1.In-house Data Yield
Idatha eveliswe kwiiseli ze-CCS ezingama-63 (ukusuka kwiintlobo ezingama-26)
| IDATHA-PacBio-CCS-15 KB | I-avareji | Max | Min | UmMedi |
| Yield - subreads (Gb) | 421.12 | 544.27 | 221.38 | 426.58 |
| Yiled - CCS(Gb) | 25.93 | 38.59 | 10.86 | 25.43 |
| Ipolymerase N50 | 145,651 | 175,430 | 118,118 | 144,689 |
| Izifundo ezingaphantsi kwe-N50 | 17,509 | 23,924 | 12,485 | 17,584 |
| I-CCS N50 | 14,490 | 19,034 | 9,876 | 14,747 |
| I-avareji yobude-Polymerase | 67,995 | 89,379 | 49,664 | 66,433 |
| I-avareji yobude-IiSubreads | 15,866 | 21,036 | 11,657 | 16,012 |
| Ubude obuphakathi-CCS | 14,489 | 19,074 | 8,575 | 14,655 |
Idatha eveliswe kwiiseli ze-CLR ze-16 (ukusuka kwiintlobo ze-76)
| IDATHA-PacBio-CLR-30Kb | I-avareji | Max | Min | UmMedi |
| Yield - subreads (Gb) | 142.20 | 291.40 | 50.55 | 142.49 |
| Ipolymerase N50 | 39,456 | 121,191 | 15,389 | 35,231 |
| Izifundo ezingaphantsi kwe-N50 | 28,490 | 41,012 | 14,430 | 29,063 |
| I-avareji yobude-Polymerase | 22,063 | 48,886 | 8,747 | 21,555 |
| I-avareji yobude-IiSubreads | 17,720 | 27,225 | 8,293 | 17,779 |
2.Idatha ye-QC - IdemoIinkcukacha-manani ngesivuno sedatha
| Isampulu | ccs Ufunda iNum | Iyonke iziseko ze-ccs (bp) | I-ccs ifunda i-N50 (bp) | ccs Ubude obuBalisiweyo (bp) | I-ccs ifundeka ixesha elide (bp) | Iziseko zofundo olungaphantsi (bp) | ireyithi ye-ccs(%) |
| PB_BMKxxx | 3,444,159 | 54,164,122,586 | 15,728 | 15,726 | 36,110 | 863,326,330,465 | 6.27 |
