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条形pasia-03

Awọn ọja

DNA/RNA lesese -PacBio Sequencer

Syeed ipasẹ PacBio jẹ ipilẹ itẹlera kika gigun, eyiti o tun mọ bi ọkan ninu awọn imọ-ẹrọ Sequencing-Iran Kẹta (TGS).Imọ-ẹrọ mojuto, akoko gidi-molecule nikan (SMRT), n fun irandiran ti kika pẹlu mewa ti kilo-mimọ ni ipari.Lori ipilẹ ti “Sequencing-by-Synthesis”, ipinnu nucleotide ẹyọkan jẹ aṣeyọri nipasẹ ipo igbi Zero-mode(ZMW), nibiti iwọn didun ti o lopin nikan ni isalẹ (ojula ti iṣelọpọ moleku), ti tan imọlẹ.Ni afikun, tito lẹsẹsẹ SMRT yago fun aibikita ọkọọkan-pato ninu eto NGS, ni pe pupọ julọ awọn igbesẹ imudara PCR ko nilo ni ilana ikole ile-ikawe.

 

Platform: Atẹle II, Revio


Awọn alaye Iṣẹ

Awọn abajade Ririnkiri

Awọn ẹya ara ẹrọ

Awọn ipo itọsẹ meji lori atẹle PacBio: Kika Gigun Tesiwaju (CLR) ati Ipinnu Ijẹwọgba Ipin (CCS)

Ipo Atẹle Iwon ikawe Data o tumq siIkore (Ni ẹẹẹkeji) Nikan-mimọYiye Awọn ohun elo
CLR 20Kb, 30Kb, ati bẹbẹ lọ. 80 Gb si 130 GB Isunmọ.85% De novo, SV pipe, ati be be lo.
CCS 15-20 Kb

14 si 40 Gb/Sẹẹli (Atẹle II)

70 si 110 Gb/Sẹli (Revio)

Da lori awọn ayẹwo

Isunmọ.99% De novo, SNP/Indel/SV pipe, Iso-Seq,

Lafiwe ti iṣẹ ati awọn ẹya ara ẹrọ ti Revio ati Sequel II

Awọn ofin

Atele II eto

Revio eto

Alekun

Ti o ga iwuwo

8 million ZMWs

25 million ZMWs

3x

Awọn ipele ominira

1

4

4x

Awọn akoko ṣiṣe kukuru

30 wakati

24 wakati

1.25x

30X HiFi eda eniyan genomes / odun

88

1.300

15x lapapọ

Awọn anfani Iṣẹ

● Lori 8 years iriri lori PacBio sequencing Syeed pẹlu egbegberun ti titi ise agbese pẹlu orisirisi eya.

● Ni ipese ni kikun pẹlu awọn iru ẹrọ PacBio Sequencing tuntun, Revio lati ṣe iṣeduro iṣelọpọ ilana ilana to to.

● Yiyara akoko titan-yika, ikore data ti o ga julọ ati data deede diẹ sii.

● Ti ṣe alabapin ninu awọn ọgọọgọrun awọn atẹjade ti o da lori ipa-giga ti PacBio.

Awọn ibeere apẹẹrẹ


Apeere Iru Iye Ifojusi (Qubit®) Iwọn didun Mimo Awọn miiran
DNA jinomiki Da lori Data ibeere ≥50 ng/μl ≥15μl OD260/280 = 1.7-2.2;
OD260/230=1.8-2.5;
Ko tente oke ni 260 nmko si kontaminesonu
Ifojusi nilo lati ṣe iwọn nipasẹ Qubit ati Qubit/Nanopore = 0.8-2.5
Apapọ RNA ≥1.2μg ≥120 ng/μl ≥15μl OD260 / 280 = 1.7-2.5;
OD260/230=0.5-2.5;ko si kontaminesonu

RIN iye ≥7.5

5≥28S/18S≥1

 

Ṣiṣẹ Iṣẹ Iṣẹ

igbaradi ayẹwo

Apeere igbaradi

Igbaradi Library

Ikole ìkàwé

Titele

Titele

Itupalẹ data

Itupalẹ data

Ayẹwo QC

Ifijiṣẹ ise agbese


  • Ti tẹlẹ:
  • Itele:

  • 1.In-house Data Ikore

    Data ti ipilẹṣẹ lati awọn sẹẹli 63 CCS (lati awọn ẹya 26)

    DATA-PacBio-CCS-15 Kb Apapọ O pọju Min Agbedemeji
    Ikore - awọn ikawe (Gb) 421.12 544.27 221.38 426.58
    Yiled - CCS(Gb) 25.93 38.59 10.86 25.43
    Polymerase N50 145.651 175.430 118.118 144,689
    Subreads N50 17.509 23,924 12.485 17.584
    CCS N50 14.490 19.034 9,876 14.747
    Apapọ ipari-Polymerase 67,995 89.379 49.664 66.433
    Apapọ ipari-Subreads 15.866 21.036 11.657 16.012
    Apapọ ipari-CCS 14.489 19.074 8.575 14.655

    Data ti ipilẹṣẹ lati awọn sẹẹli 16 CLR (lati awọn ẹya 76)

    DATA-PacBio-CLR-30Kb Apapọ O pọju Min Agbedemeji
    Ikore - awọn ikawe (Gb) 142.20 291.40 50.55 142.49
    Polymerase N50 39.456 121.191 15.389 35.231
    Subreads N50 28.490 41.012 14.430 29.063
    Apapọ ipari-Polymerase 22.063 48.886 8.747 21.555
    Apapọ ipari-Subreads 17.720 27.225 8.293 17.779

    2.Data QC - RirinkiriAwọn iṣiro lori ikore data

    Apeere

    ccs Say Nọm

    Lapapọ Awọn ipilẹ ccs (bp)

    ccs Ka N50 (bp)

    Itumọ Gigun (bp)

    ccs kika ti o gun julọ (bp)

    Awọn ipilẹ abẹlẹ (bp)

    oṣuwọn ccs (%)

    PB_BMKxxx

    3,444,159

    54.164.122.586

    15.728

    15.726

    36,110

    863,326,330,465

    6.27

     

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