DNA / RNA Sequencing -Pacbio
EGO
Usoro ihe abụọ na-adịghị mma na pacbio sequencer: Gụ na-agụ (CLR) ma na-agụ ihe okirikiri (CCS)
| Ọnọdụ Sequencing | Ọbá akwụkwọ | Akwụkwọ NsọMkpụrụ (kwa sel) | Otu-ntọalaKpomkwem | Ngwa |
| Nka | 20KB, 30KB, wdg. | 80 GB TO 130 GB | Odika. 85% | De novo, SV na-akpọ, wdg. |
| CCS | 15-20 kb | 14 ruo 40 GB / Cell (Sequel II) 70 ruo 110 GB / Cell (Revio) Dabere na nlele | Odika. 99% | De novo, SNP / INV / SV kpọrọ, ISO-SOQ, |
Ntụnyere nke arụmọrụ na atụmatụ nke Revio na sequel II
| Okwu | Sistel II | Usoro revoo | Mmubanye |
| Nnọọ dị elu | 8 Nde ZMWs | 25 nde ZMWs | 3x |
| Ohere dị iche iche | 1 | 4 | 4x |
| Oge ịgba ọsọ | 30 awa | 24 awa | 1.25x |
| 30x HIFI Gonimes / Afọ | 88 | 1,300 | 15X niile |
Uru bara uru
● Ihe karịrị afọ asatọ na-ahụ na Pacbio Sequiving nwere puku kwuru puku ndị otu emechiri emechi.
Jiri nyiwe zuru oke nke Pacbio na-eme zuru ezu, nlọtọ na-ekwe nkwa inweta ntozu oke.
● Ntughari ngwa ngwa, elu data dị elu na data ziri ezi.
● Enyere onyinye na ọtụtụ narị mbipụta Pacbio.
Nlele Nyere
| Ụdị atụ | Ego | Ileta (Qubit®) | Ibu | Nko | Ndi ozo |
| Gnamic DNA | Dabere na ihe achọrọ | ≥50 NG / μl | ≥15μl | OD260 / 280 = 1.7-2.2; OD260 / 230 = 1.8-2.5; O doro anya na 260 NM,Ọ dịghị ihe mgbochi | Izigide chọrọ ka a tụọ ya na Qubit / Nanopore = 0.8-2.5 |
| Ngụkọta rna | ≥1.2μμG | ≥120 NG / μl | ≥15μl | OD260 / 280 = 1.7-2.5; OD260 / 230 = 0.5-2.5;Ọ dịghị ihe mgbochi | Rin Uru ≥7.5 5≥28s / 18s≥1 |
Ọrụ Ọrụ
Nkwadebe nlele
Ọbá Akwụkwọ Ewu
Sequencing
Nyocha data
Nnyefe oru ngo
Mkpụrụ data 1.in-ụlọ
Data sitere na mkpụrụ ndụ 63 CCS (site na ụdị 26)
| Data-Pacbio-ccs-15 KB | Agbaetiti | Marnet | Mine | Median |
| Mkpụrụ - Sperreads (GB) | 421.12 | 544.27 | 221.38 | 426.58 |
| Yiled - CCS (GB) | 25.93 | 38.59 | 10.86 | 25.43 |
| Polmerase N50 | 145,651 | 175,430 | 118,118 | 144,689 |
| Ndenye N50 | 17,509 | 23,924 | 12,485 | 17,584 |
| CCS N50 | 14,490 | 19,034 | 9,876 | 14,747 |
| Nkezi ogologo-polymerase | 67,995 | 89,379 | 49,664 | 66,433 |
| Nkezi ogologo-subraads | 15,866 | 21,036 | 11,657 | 16,012 |
| Nkezi ogologo-ccs | 14,489 | 19,074 | 8,575 | 14,655 |
Data site na mkpụrụ ndụ 16 CLR (site na ụdị 76)
| Data-pacbio-crr-30kb | Agbaetiti | Marnet | Mine | Median |
| Mkpụrụ - Sperreads (GB) | 142.20 | 291.40 | 50.55 | 142.49 |
| Polmerase N50 | 39,456 | 121,191 | 15,389 | 35,231 |
| Ndenye N50 | 28,490 | 41,012 | 14,430 | 29,063 |
| Nkezi ogologo-polymerase | 22,063 | 48,886 | 8,747 | 21,5555 |
| Nkezi ogologo-subraads | 17,720 | 27,225 | 8,293 | 17,779 |
2.Data QC - ngosiNdekọ na mkpụrụ data
| Sampul | CCS na-agụ nọmba | Ngụkọta ntọala CCS (BP) | CCS na-agụ N50 (BP) | CCS pụtara ogologo (BP) | ccs ogologo oge (BP) | Ntọala dị n'okpuru (BP) | Ọnụego CCS (%) |
| Pb_bmkxxx | 3,444,159 | 54,164,122,586 | 15,728 | 15,726 | 36,110 | 863,326,330,465 | 6.27 |
