Yakazara-kureba mRNA kutevedzana -PacBio
Features
● cDNA synthesis kubva ku poly-A mRNA inoteverwa nekugadzirira raibhurari
● Sequencing muCCS mode, kugadzira HiFi kuverenga
● Kutevedzana kwezvinyorwa zvehurefu hwakazara
● Kuongorora hakudi referensi genome; zvisinei, inogona kushandiswa
● Kuongorora kweBioinformamatic kunogonesa kuongororwa kwezvinyorwa isoform lncRNA, gene fusions, poly-adenylation, uye gene chimiro.
Basa Rinobatsira
●Kururama Kwepamusoro: HiFi inoverenga nemazvo> 99.9% (Q30), inofananidzwa neNGS
● Alternative Splicing Analysis: kutevedzana kwezvinyorwa zvese zvinogonesa isoform kuzivikanwa uye hunhu
●Hunyanzvi Hwakawanda: iine rekodhi yekupedzisa anopfuura zana negumi nemazana ePacBio akazara-kureba transcriptome mapurojekiti uye kugadzirisa anopfuura mazana maviri nemakumi matatu emasampuli, timu yedu inounza hupfumi hweruzivo kupurojekiti yega yega.
●Post-Sales Support: kuzvipira kwedu kunowedzera kupfuura kupera kweprojekiti nemwedzi mitatu mushure mekutengesa sevhisi nguva. Munguva ino, tinopa kutevedza kweprojekiti, rubatsiro rwekugadzirisa matambudziko, uye maQ&A masesheni kugadzirisa chero mibvunzo ine chekuita nemhedzisiro.
Sample Zvinodiwa uye Delivery
| Library | Sequencing strategy | Data inokurudzirwa | Quality Control |
| PolyA yakapfumisa mRNA CCS raibhurari | PacBio Sequel II PacBio Revio | 20/40 Gb 5/10 M CCS | Q30≥85% |
Sample Zvinodiwa:
Nucleotides:
● Zvirimwa:
Mudzi, Dzinde kana Petal: 450 mg
Shizha kana Mbeu: 300 mg
Muchero: 1.2 g
● Mhuka:
Mwoyo kana Ura: 300 mg
Viscera kana Brain: 240 mg
Tsandanyama: 450 mg
Mabhonzo, Bvudzi kana Ganda: 1g
● Arthropods:
Zvipembenene: 6g
Crustacea: 300 mg
● Ropa rose: 1 chubhu
● Masero: 106 masero
| Conc.(ng/μl) | Mari (μg) | Kuchena | Kuperera |
| ≥ 100 | ≥ 1.0 | OD260/280=1.7-2.5 OD260/230=0.5-2.5 Yakaganhurirwa kana isina mapuroteni kana kusvibiswa kweDNA kunoratidzwa pagel. | Kumiti: RIN≥7.5; Zvemhuka: RIN≥8.0; 5.0≥ 28S/18S≥1.0; kukwirira kwakadzika kana kusina kwekutanga |
Yakakurudzirwa Sample Delivery
Container: 2 ml centrifuge chubhu (Tin foil haina kukurudzirwa)
Sample labeling: Boka+ replicate eg A1, A2, A3; B1, B2, B3.
Shipment:
1. Dry-ice: Samples dzinoda kurongedzerwa mumabhegi uye kuvigwa muoma-aizi.
2. RNAstable tubes: RNA samples inogona kuomeswa muRNA stabilization tube (eg RNAstable®) uye inotumirwa mukamuri yekupisa.
Kuyerera Kwebasa Rebasa
Kuedza kugadzira
Muenzaniso wekutumira
RNA kubviswa
Kuvakwa kweraibhurari
Sequencing
Data analysis
Mushure mekutengesa masevhisi
Inosanganisira ongororo inotevera:
● Raw data quality control
● Alternative Polyadenylation Analysis (APA)
● Fusion chinyorwa chekuongorora
● Alternative Splicing Analysis
● Benchmarking Universal Single-Copy Orthologs (BUSCO) kuongorora
● Novel transcript analysis: kufanotaura kwekodhi sequences (CDS) uye annotation inoshanda
● lncRNA kuongorora: kufanotaura kwelncRNA uye zvinangwa
● MicroSatelite Identification (SSR)
BUSCO ongororo
Alternative Splicing Analysis
Alternative Polyadenylation Analysis (APA)
Tsanangudzo inoshanda yezvinyorwa zvitsva
Ongorora kufambira mberi kwakafambiswa neBMKGene's Nanopore yakazara-kureba mRNA sequencing masevhisi mune ino chinyorwa.
Ma, Y. nevamwe. (2023) 'Kuenzanisa kuongorora kwePacBio uye ONT RNA kutevedza nzira dzeNemopilema Nomurai chepfu identification', Genomics, 115 (6), p. 110709. doi: 10.1016/J.YGENO.2023.110709.
Chao, Q. et al. (2019) 'Masimba ekusimudzira ePopulus stem transcriptome', Plant Biotechnology Journal, 17 (1), mapeji 206-219. doi: 10.1111/PBI.12958.
Deng, H. et al. (2022) 'Dynamic Changes in Ascorbic Acid Content during Fruit Development and Ripening of Actinidia latifolia (an Ascorbate-Rich Fruit Crop) uye Associated Molecular Mechanisms', International Journal of Molecular Sciences, 23 (10), p. 5808. doi: 10.3390/IJMS23105808/S1.
Hua, X. nevamwe. (2022) 'Kufanotaura kunobudirira kwemajini enzira yebiosynthetic inobatanidzwa mubioactive polyphyllins muParis polyphylla', Communications Biology 2022 5: 1, 5 (1), mapeji 1-10. doi: 10.1038/s42003-022-03000-z.
Liu, M. nevamwe. (2023) 'Yakasanganiswa PacBio Iso-Seq uye Illumina RNA-Seq Kuongorora kweTuta absoluta (Meyrick) Transcriptome uye Cytochrome P450 Genes', Zvipembenene, 14 (4), p. 363. doi: 10.3390/INSECTS14040363/S1.
Wang, Lijun et al. (2019) 'Ongororo ye transcriptome yakaoma uchishandisa PacBio single-molecule chaiyo-nguva yekuongorora yakasanganiswa neIllumina RNA sequencing yekunzwisisa zviri nani ricinoleic acid biosynthesis muRicinus communis', BMC Genomics, 20 (1), mapeji 1-17. doi: 10.1186/S12864-019-5832-9.
