PERBANDINGAN TIGA KIT EKSTRAKSI RNA UNTUK ANALISIS TRANSKRIPTOMIKA PADA KELAPA SAWIT (Elaeis guineensis Jacq.)
Main Article Content
Abstract
Menghasilkan RNA berkualitas tinggi sangatlah penting pada tahap awal penelitian biologi molekuler. Untuk mengisolasi RNA diperlukan keterampilan dan kehati-hatian tinggi dalam mengikuti prosedur di laboratorium karena RNA lebih mudah terdegradasi, khususnya sampel hasil kultur jaringan tanaman. Salah satu parameter yang digunakan pada pengecekan kualitas RNA total adalah RIN (RNA Integrity Number). Penelitian bertujuan mendapatkan metode ekstraksi RNA pada daun, kalus dan embrio somatik kelapa sawit yang berkualitas baik dan memiliki konsentrasi tinggi untuk analisa transkriptomika. Ekstraksi RNA dilakukan menggunakan metode kit Plant RNA PureLink (Ambion), Genezol RNA Extraction (Geneaid) dan RibospinTM Plant (Geneall). Hasil menunjukkan bahwa RNA daun, kalus dan embrio somatik kelapa sawit telah berhasil diekstraksi dengan menggunakan kit RibospinTM (Geneall). RNA hasil ekstraksi tersebut dapat digunakan untuk sekuensing RNA dengan tujuan analisis transkriptomika, dilihat dari jumlah total RNA yang lebih dari 4 μg dan nilai RIN lebih dari 7.
Article Details
This work is licensed under a Creative Commons Attribution-NonCommercial-ShareAlike 4.0 International License.
Authors who publish with this journal agree to the following terms:
a). Authors retain copyright and grant the journal right of first publication with the work simultaneously licensed under a Attribution-NonCommercial-ShareAlike 4.0 International that allows others to share the work with an acknowledgement of the work's authorship and initial publication in this journal.
b). Authors are able to enter into separate, additional contractual arrangements for the non-exclusive distribution of the journal's published version of the work (e.g., post it to an institutional repository or publish it in a book), with an acknowledgement of its initial publication in this journal.
c). Authors are permitted and encouraged to post their work online (e.g., in institutional repositories or on their website) prior to and during the submission process, as it can lead to productive exchanges, as well as earlier and greater citation of published work (See The Effect of Open Access).
References
Agrawal AA (2011) Current trends in the evolutionary ecology of plant defence. Funct Ecol 25:420-432. doi: 10.1111/j.1365-2435.2010.01796.x
Ambion (2012) PureLink® RNA Mini Kit handbook. Carlsbad, CA USA. https://www.thermofisher.com/document-connect/documentconnect.html?url= https://assets.thermofisher.com/TFSAssets/LSG/manuals/purelink_rna_mini_kit_man.pdf&title=UHVyZUxpbmsgUk5BIE1pbmkgS2l0. Diakses 22 Februari 2019
Aranda R, Dineen SM, Craig RL, Guerrieri RA, Robertson JM (2009) Comparison and evaluation of RNA quantification methods using viral, prokaryotic, and eukaryotic RNA over a 10 4 concentration range. Anal Biochem 387:122-127. doi: 10.1016/j.ab.2009.01.003
Becker C, Hammerle-Fickinger A, Riedmaier I, Pfaffl MW (2010) mRNA and microRNA quality control for RT-qPCR analysis. Methods 50:237-243. doi: 10.1016/j.ymeth.2010.01.010
Berenbaum MR, Zangerl AR (2008) Facing the future of plant-insect interaction research: le retour a la ‘‘Raison d’Etre’’. Plant Physiol 146:804-811. doi: 10.1104/pp.107.113472
Bilgin DD, DeLucia EH, Clough SJ (2009) A robust plant RNA isolation method suitable for Affymetrix GeneChip analysis and quantitative real-time RT-PCR. Nat Protoc 4:333-340. doi: 10.1038/nprot.2008.249
Briones C, Nuñez JJ, Pérez M, Espinoza-Rojas D, Molina-Quiroz C, Guiñez R (2018) De novo male gonad transcriptome draft for the marine mussel Perumytilus purpuratus with a focus on its reproductive-related proteins. J Genomics 6:127-132. doi: 10.7150/jgen.27864
Bryant S, Manning DL (1998) Isolation of messenger RNA. Methods Mol Biol 86:61-64. doi: 10.1385/0-89603-494-1:61
Daohong W, Bochu W, Biao L, Chuanren D, Jin Z (2004) Extraction of total RNA from Chrysanthemum containing high levels of phenolic and carbohydrates. Colloids Surfaces B Biointerfaces 36:111-114. doi: 10.1016/J.COLSURFB.2004.05.014
Farrell RE (2010) RNA Methodologies: A laboratory guide for isolation and characterization. 4th Edition. Academic Press, London. doi: 10.1016/C2009-0-01850-9
Fleige S, Walf V, Huch S, Prgomet C, Sehm J, Pfaffl MW (2006) Comparison of relative mRNA quantification models and the impact of RNA integrity in quantitative real-time RT-PCR. Biotechnol Lett 28:1601-163. doi: 10.1007/s10529-006-9127-2
Gallego Romero I, Pai AA, Tung J, Gilad Y (2014) RNA-seq: impact of RNA degradation on transcript quantification. BMC Biology 12:42 doi: 10.1186/1741-7007-12-42
Gayral P, Weinert L, Chiari Y, Tsagkogeorga G, Ballenghien M, Galtier N (2011) Next-generation sequencing of transcriptomes: A guide to RNA isolation in nonmodel animals Mol Ecol Res 11:650-661. doi: 10.1111/j.1755-0998.2011.03010.x
Geneaid (2015) GENEzol™ Reagent handbook, Taiwan. http://www.geneaid.com/sites/default/files/GZR8.pdf. Diakses 22 Februari 2019
GeneAll (2012) GeneAll® RibospinTM Plant. Plant total RNA purification Handbook. Brief protocol. http://www.tribioscience.com/files/307-150.pdf. Diakses 22 Februari 2019
Habib SH, Saud HM, Kausar H (2014) Efficient oil palm total RNA extraction with a total RNA extraction kit. Genet Mol Res 13:2359-2367 doi: 10.4238/201
Jeffries MKS, Kiss AJ, Smith AW, Oris JT (2014) A comparison of commercially-available automated and manual extraction kits for the isolation of total RNA from small tissue samples. BMC Biotechnol 14:94. doi: 10.1186/s12896-014-0094-8
Kiewe P, Gueller S, Komor M, Stroux A, Theil E, Hoffman WK (2009) Prediction of qualitative outcome of oligonucleotide microarray hybridization by measurement of RNA integrity using the 2100 Bioanalyzer capillary electrophoresis system Ann Hematol 88:1177-1183. doi: 10.1007/s00277-009-0751-5
Macel M, van Dam NM, Keurentjes JJB (2010) Metabolomics: The chemistry between ecology and genetics. Mol Ecol Resour 10:583-593. doi: 10.1111/j.1755-0998.2010.02854.x
Mueller O, Ligthfoot S, Schroeder A (2016) RNA Integrity Number (RIN): Standardization of RNA quality control application. Agilent application note publication 1:1-8 https://www.agilent.com/cs/library/applications/5989-1165EN.pdf. Diakses 22 Februari 2019
Opitz L, Salinas-Riester G, Grade M, Jung K, Jo P, Emons G, Ghadimi BM, Beißbarth T, Gaedcke J (2010) Impact of RNA degradation on gene expression profiling. BMC Med Genomics 3:36. doi: 10.1186/1755-8794-3-36
Pereira WJ, Bassinello PZ, Brondani C, Vianello RP (2017) An improved method for RNA extraction from common bean seeds and validation of reference genes for qPCR. Crop Breed Appl Biotechnol 17:150-158. doi: 10.1590/1984-70332017v17n2a22
Qadri R, Iqbal A, Wu Y, Li J, Nisar N, Azam M, Yang Y (2019) A modified protocol for total RNA isolation from different oil palm (Elaeis guineensis) tissues using cetyltrimethylammonium bromide. Curr Sci 116:479-482 doi: 10.18520/cs/v116/i3/479-482
Qiao G, Liu M, Song K, Li H, Yang H, Yin Y, Zhuo R (2017) Phenotypic and comparative transcriptome analysis of different ploidy plants in Dendrocalamus latiflorus Munro. Front Plant Sci 8:1371 doi: 10.3389/fpls.2017.01371
Reiman M, Laan M, Rull K, Sõber S (2017) Effects of RNA integrity on transcript quantification by total RNA sequencing of clinically collected human placental samples FASEB J 31:3298-3308. doi: 10.1096/fj.201601031RR
Sambrook J, Russell D (2001) Molecular cloning: A laboratory manual. 3rd edition. Cold Spring Harbor Laboratory Press, New York
Schroeder A, Mueller O, Stocker S, Salowsky R, Leiber M, Gassmann M, Ligthfoot S, Menzel W, Granzow M, Ragg T (2006) The RIN: an RNA integrity number for assigning integrity value to RNA measurements. BMC Mol Biol 7:3. doi: 10.1186/1471-2199-7-3
Tesena P, Korchunjit W, Taylor J, Wongtawan T (2017) Comparison of commercial RNA extraction kits and qPCR master mixes for studying gene expression in small biopsy tissue samples from the equine gastric epithelium. J Equine Sci 28:135-141. doi: 10.1294/jes.28.135.
Walter J, Hein R, Auge H, Beierkuhnlein C, Loeffler S (2012) How do extreme drought and plant community composition affect host plant metabolites and herbivore performance? Arthropod-Plant Interact 6:15-25. doi: 10.1007/s11829-011-9157-0
Xiao Y, Yang Y, Cao H, Fan H, Ma Z, Lei X, Mason AS, Xia Z, Huang X (2012) Efficient isolation of high quality RNA from tropical palms for RNA-seq analysis. Plant Omics 5:584-589