01 sirna AccuTarget TM sirnas 35 Custom sirna 36 Genome-wide sirna 38 AccuTarget Genome-wide Predesigned sirna Library 39 AccuTarget Premade sirna Set

Transcription

1 Custom sirna Genome-wide sirna sirna Oligonucleotide FAQs AccuTarget mirna mimics & inhibitors AccuTarget mirna Controls sirna & mirna Phone: (ext.4->1)

2 01 sirna AccuTarget TM sirnas 35 Custom sirna 36 Genome-wide sirna 38 AccuTarget Genome-wide Predesigned sirna Library 39 AccuTarget Premade sirna Sets 40 AccuTarget Control sirnas 42 AccuTarget Real-Time PCR Primers Library 46 sirna Oligonucleotide FAQs 48 User Protocol 49

3 AccuTarget TM sirnas Overview sirna(small interference RNA) 는 target specific gene silencing 을위한가장강력한수단으로유전자기능의확인, drug target 의 validation, 각종질병의 gene therapy 등에사용되고있습니다. sirna 는기존 antisense technology 에비해다음과같은장점을가지고있습니다. - 첫째, 적은수의 screening 으로도높은효율의 sirna 를찾을수있어시간과경비를절감할수있습니다. - 둘째, 낮은농도로효율적인 gene silencing 이가능합니다. - 셋째, natural biological mechanism 으로기작이매우 target specific 합니다. sirna mechanism sirna 는약 21-nucleotide sense 와 anti-sense RNA 로구성된 double-strand RNA 입니다. 이들이세포안으로전달되면 sirna 는 RNA-induced silencing complex(risc) 라고불리는 protein complex 와결합합니다. single strand RNA 가 RISC 에의해선택및장착되어 ribonucleoprotein complex 를형성하고, 이러한과정을통하여활성화된 RISC 의 single strand RNA 는상보적인서열의 mrna target 에결합하여 target mrna 를분해합니다. 35

4 Custom sirna Description Custom sirna 는고객이요구하는 sirna sequence 를합성해드리는서비스입니다. 바이오니아에서 sirna 를합성하는경우에는 free-design 서비스를제공받을수있습니다. 본사의 AccuTarget TM Genome-wide Pre-designed sirna 는확인해보시면, Human ( 약 18,048 개 ), Mouse ( 약 17,117 개 ) 와 Rat ( 약 9,392 개 ) 의 pre-designed sirna 를합성하여즉시배송가능한제품입니다. Pre-designed sirna 이외의유전자는 Turbo si- Designer 에의해제작및합성이가능합니다. 또한하나의유전자에서세개이상의 sirna design 및합성도가능합니다. sirna 정제방법으로 Bio-RP, HPLC 정제를선택하실수있습니다. 또한원하시는 sirna sequence 에다양한 modification 도가능합니다. 30 mer 의 sirna 를최대 32 개의서로다른 3 overhang 선택을포함한다양한옵션을선택하여다양한형태로 sirna 를주문할수있습니다. sirna 는정제방법, annealing 유 / 무, dry 또는 ready-to-use 등의선택옵션에따라제작되어제공됩니다. Figure 1. MALDI-TOF mass spectrometry analysis of a custom sirna. All sirnas are subjected to MALDI-TOF mass spectrometry to ensure its quality. Modification 문의는 sirna-support@bioneer.co.kr 로메일을보내주시거나 042) 으로연락주시기바랍니다. 모든 Custom sirna 는본사의최첨단 clean room 시설에서합성된후바이오니아의 Bio-RP purification system 을통해정제됩니다. 생산된모든 sirna 는 MALDI-TOF mass spectrometry 를이용하여분석하고, 합격된 sirna 만을발송합니다. 보다높은순도의 sirna 를원하신다면, HPLC purification 선택을추천합니다 (Figure 1). Annealed sirna 를요청하신경우에는동량의 sense 와 antisense RNA 를 annealing 하여 double strand form 으로제작한다음, PAGE 로 double strand sirna 가제대로만들어졌는지확인한후제공해드립니다 (Figure 2). Features and Benefits Guaranteed performance: 3 개중 2 개의 custom sirna 80% 이상의 knockdown 효율을보입니다. AccuTarget TM sirna 는 3 개중 2 개가적어도 80% 의 knockdown 효율을보입니다, 만약그렇지않을경우무료로 2 개의 sirna 를추가로제공합니다. Guaranteed quality: 모든 sirna 는 Clean room 에서제조되며 MALDI-TOF 및 PAGE 로검증됩니다 Custom sirna design service available: 무료로 Turbo si-designer software 를사용하여무료로제작됩니다. Competitive pricing: 무료로 Overhang 서비스가제공되므로경제적비용으로제품을이용할수있습니다. Figure 2. PAGE data of annealed double-stranded custom sirna. Complementary single-strand RNA strands were hybridized to form sirna duplex and analyzed by 15% non-denaturing PAGE. SS: single-strand RNA, DS: double-strand sirna. Ordering Information Synthesis/Purification Price Cat. no. S Purification Guaranteed nmole Price ( 원 ) 10 nmole 120,000 S nmole 145,000 Bio-RP S nmole 230,000 S nmole 330,000 S nmole 165,000 S nmole 240,000 HPLC S nmole 380,000 S nmole 540,000 Single strand 형태의 sense 및 antisense RNA 각각을동일한농도로 dry 하여제공해드립니다. Annealing buffer 와 DEPC D.W. 를기본으로제공해드립니다. Annealing 된상태의 sirna 를원하실경우는 2 만원의비용이추가됩니다. 모든합성 RNA 에대해 MALDI-TOF QC data 를첨부해드립니다. BIONEER 36

5 Custom sirna Modification Oligos Price 5 - Modification 단위 ( 원 ) Product Purification 10 nmole 20 nmole 50 nmole 100 nmole 5' Fluorescein 70,000 90, , ,000 5' Phosphorylation 22,000 30,000 41,000 53,000 5' Biotin 41,000 50,000 73, ,000 5' Amine 45,000 81, , ,000 5' TAMRA 135, , , ,000 HPLC 5' Thiol 125, , , ,000 5' Cy3 180, , , ,000 5' Cy5 180, , , ,000 5' PEG 2000 Inquire Inquire Inquire Inquire 5 Cy , , , , Modification 단위 ( 원 ) Product Purification 10 nmole 20 nmole 50 nmole 100 nmole 3' Fluorescein 80, , , ,000 3' Phosphorylation 60, , , ,000 3' Biotin 130, , , ,000 3' Amine 60, , , ,000 3' TAMRA 135, , , ,000 HPLC 3' Thiol 125, , , ,000 3' DABCYL 250, , , ,000 3' Cholesterol 130, , , ,000 3' PEG 2000 Inquire Inquire Inquire Inquire 3 Cy , , , ,000 Internal Modification 단위 ( 원 ) Product Purification 10 nmole 20 nmole 50 nmole 100 nmole Phosphorothioate - 5,500 9,000 12,000 2'-OMe-rA, rc, rg, ru 8,500 11,000 16,000 22,000 2'-F(A) 28,000 32,000 38,000 50,000 2'-F(U) 28,000 32,000 38,000 50,000 2'-F(G) HPLC 28,000 32,000 38,000 50,000 2'-F(C) 28,000 32,000 38,000 50,000 Inosine 17,000 19,000 25,000 31,000 Deoxy-abase 49,000 58,000 70,000 94,000 Chimeric DNA 3,500 5,000 7,500 12,000 그외 Modification 합성여부및가격은별도문의바랍니다. 신속하고전문적인고객서비스 sirna 실험이익숙하지않은고객들도본사의제품을구입하시면언제든지실험관련상담을받으실수있습니다. sirna 를이용한다양한실험들에대해수년간의경험이축적된바이오니아의연구원들이고객들의질문에대해신속하고친절하게답해드립니다. 제품및실험관련문의사항은아래이메일또는전화번호로연락을주시면당일에성실히답변해드립니다. sirna 제품및실험관련상담 sirna@bioneer.co.kr, sirna-support@bioneer.co.kr Tel: ( 올리고팀 ) 37

6 Genome-wide sirna Overview Turbo si-designer - Bioneer s proprietary sirna design algorithm Small interfering RNA(siRNA) 는최근새로운 functional genomics 의 tool 로서각광을받고있고, 그중요성을인정받아 2002 년도 Science 저널에서 small RNA(siRNA 와 mirna) 가 Breakthrough of the year 1 번으로선정되었습니다. 이와같은 RNAi 연구에서가장중요한요소가운데하나는 효율이높은 target site 를찾는가 입니다. Target site 마다이에상응하는 sirna 의효율이다르기때문에, 고효율의 sirna target site 를찾는것이분명한실험결과를얻는데매우중요합니다. 바이오니아에서는 2003 년 10 월부터산업자원부지원하에 sirna Design algorithm 개발프로젝트를시작하였고주관기관인 ( 주 ) 바이오니아와위탁기관인국가유전체정보센터 (NGIC, 책임자 ; 김영주박사 ) 가공동으로 3 년에걸쳐총 9 억원을투입하여효능이뛰어난 sirna design algorithm 을개발하였습니다. 본과제의주목적은기존의연구를통해이미알려져있는 nonfunctional 한 sirna site 를배제시키고다른여러변수들을고려하여효율적인 sirna target site 를찾는 algorithm 을개발하는것이었습니다. 이렇게개발된일차적인 algorithm 을이용하여예측한 sirna target site 를실험을통해그효과를확인하고, 각변수가실험결과에미치는영향을분석하여보다효율적인 sirna target site 를높은확률로예측할수있는 design program 을만들어내었습니다. Turbo si-designer 의성능을확인하기위하여항세포사멸및세포분열관련유전자인 Survivin 을대상으로 82 개의 sirna 를 design 하여 A549 lung carcinoma 세포에 transfection 후, Northern Blot 과 QuantiGene kit 로 mrna 의 knockdown 을측정하였습니다. Figure. 1B 에서보듯이점수가낮은 sirna 들은점수가높은 Figure. 1A 에비해효율이좋지않음을알수있고따라서 Turbo si-designer 가비효율적인 sirna site 들을매우효과적으로배제함으로써고효율의 sirna 를찾을확률을높여주는것을알수있습니다.(Figure 1) Figure 1. sirna knockdown efficiency of sirnas designed by Turbo si-designer was analyzed by Northern blot and Real-Time PCR analysis. A) sirna knockdown efficiency of high NGIC score sirnas. B) sirna knockdown efficiency of low NGIC score sirnas BIONEER 38

7 AccuTarget Genome-wide Predesigned sirna Library Description AccuTarget TM Genome-wide predesigned sirna 는한국생명공학연구원과바이오니아가공동으로개발한 sirna design algorithm 인 Turbo si-designer 를이용하여 predesign 된 sirna 입니다. 이를사용하여 knockdown 효율이우수한 AccuTarget TM Genome-wide predesigned sirna database 를구축하였습니다. The Bioneer Guarantee 같은유전자에대한 3 개의 sirna 를구입하시면, 3 개의 sirna 중 2 개가 target mrna expression level 을 80% 이상감소시킬것을보장합니다. Target 유전자의 mrna 수준에서 80% 이상의감소가없을경우, 무료로 2 개의 sirna 를제공합니다. * 단, 본사에서요하는다음과같은 supporting data 를별도로제출하셔 야합니다. 1. sirna knockdown efficiency data: NC (AccuTarget TM Negative Control) and sirna concentration at 100 nm 2. Transfection efficiency data: PC (AccuTarget TM GAPDH/GFP/ Luciferase sirna) and NC (AccuTarget TM Fluorescein-labeled Negative Control) Features and Benefits High sirna knockdown rates: 주문한 3 개의 sirna 중 2 개는 target mrna expression level 을 80% 이상억제합니다. Unique design algorithm: Off target effect 를최소화하면서 sirna knockdown 효율을최대화합니다. Competitive pricing: 경제적인비용으로극대화된효율을경험하세요. Application AccuTarget TM Genome-wide Predesigned sirna Library 는다양한 RNAi 실험에활용할수있습니다. Functional genomics and proteomics research Gene expression studies Array analysis Ordering Information Purification Bio-RP HPLC Guaranteed nmole Price ( 원 ) 10 nmole 140, nmole 165, nmole 250, nmole 350, nmole 185, nmole 260, nmole 400, nmole 560,000 Figure 2. sirna knockdown efficiency of AccuTarget TM Genome-wide Predesigned sirna. AccuTarget TM Predesigned sirnas are highly effective. To determine sirna knockdown efficiency of predesigned sirnas, HeLa cells were transfected with sirnas at 100 nm concentration. 24 hr post-transfection, total RNA was isolated and the level of target mrna was measured by qrt-pcr. This data demonstrates the effectiveness of the Turbo si-designer algorithm: 83.8% of tested sirnas induced >70% sirna knockdown and 38.1% of tested sirnas elicited >90% knockdown. 39

8 AccuTarget Premade sirna Sets Description AccuTarget TM premade human sirna Library 는즉시 transfection 실험에사용할수있도록제작된 54,144 sirna 로이루어진제품입니다. Premade human sirna Library 는 10, 20, 50, 100 nmole 의다양한용량으로주문가능합니다. 이러한 Premade sirna library 에포함되어있는개별 sirna 를주문할수있으며 10, 20, 50 또는 100 nmole guaranteed yield 로주문가능합니다. 또한, 특정 cellular process, 암, 질병등의연구목적에특화된 25 개의 biological pathway ( 혹은 gene family) 별로제작된 sirna library set 를제공합니다. 이러한 library set 는각 sirna 의 guaranteed yield 0.1, 0.25, 0.5, 1 nmole 로주문가능합니다. 실제 cell culture 실험에서높은 knockdown 효능이검증된 sirna 로이루어진 21 개의 Pre-validated library 를제공합니다. 691 개의유전자에대해만들어진 Pre-validated sirna library 에포함되어있는개별 sirna 를 10, 20, 50, 또는 100 nmole guaranteed yield 로주문할수있습니다. 이렇게디자인된 sirna validation 과정은다음과같습니다. Features and Benefits Categorized by pathway / family: 연구수행에편리하게특화된 format Pre-validated sirna libraries available: 즉시사용가능한검증된효능의 sirna library Competitive pricing: 경제적인비용으로극대화된효율을경험하세요. Applications AccuTarget TM Premade sirna set 은다양한 RNAi 실험에활용할수있습니다. Pathway analysis and target identification and validation Drug target HTS (High Throughput sirna Screening) BIONEER 40

9 AccuTarget Premade sirna Sets Ordering Information Cat. no. Product Name AccuTarget TM Premade Human sirna sets No. of Genes Cat. no. Product Name No. of Genes SHS-001 Antioxidant sirna set 38 SHS-013 Lyase sirna set 123 SHS-002 Apoptosis sirna set 290 SHS-014 Motor sirna set 122 SHS-025 Cancer sirna set 1157 SHS-015 NF-kB pathway sirna set 37 SHS-003 Caspase sirna set 37 SHS-016 Nucleic acid binding sirna set 2573 SHS-004 Cell cycle sirna set 112 SHS-017 Oxidoreductase sirna set 551 SHS-005 Cyclase sirna set 21 SHS-018 Peptidase sirna set 491 SHS-006 Cytochrome P450 sirna set 52 SHS-019 Phosphatase sirna set 188 SHS-007 Deaminase sirna set 22 SHS-020 Receptor sirna set 1516 SHS-008 GPCR signaling pathway sirna set 727 SHS-021 Transferase sirna set 1428 SHS-009 Helicase sirna set 114 SHS-022 Transporter sirna set 1021 SHS-010 Isomerase sirna set 104 SHS-023 Tubulin sirna set 20 SHS-011 Kinase sirna set 699 SHS-024 Ubiquitin sirna set 77 SHS-012 Ligase sirna set 272 Figure 3. Knockdown efficiency of AccuTarget TM sirna library. To determine knockdown efficiency, HeLa cells were transfected with individual sirnas at 100 nm concentration. 24 hr post-transfection, total RNA was isolated and the level of target mrna was measured by qrt-pcr. This data demonstrates the effectiveness of the Turbo si-designer algorithm: 83.8% of tested sirnas induced >70% knockdown and 38.1% of tested sirnas elicited >90% knockdown. Figure 4. sirna knockdown efficiency of AccuTarget TM Human Validated Cell Cycle sirna set. HeLa cells were transfected with sirnas targeting 33 different genes at a concentration of 20 nm. Total RNA was isolated and the level of target mrna was measured by qrt-pcr. This data demonstrates the effectiveness of the Turbo si-designer algorithm. 41

10 AccuTarget Control sirnas Description AccuTarget TM Positive Control sirna 는 target gene 에대하여높은효율의 knockdown 효과를나타냅니다 (Figure 5, 6 & 7). 널리사용되는 housekeeping gene 인 GAPDH 에대한효율적인 sirna 를제공하며 reporter system 으로 GFP 와 Luciferase 에대한 positive control sirna 도제공합니다. 또한 Lamin A/C or PPIB(cyclophilin B) 에대해효율적인 mouse positive control sirna 를제공합니다. Human 과 mouse, rat 에서공통으로사용가능한 AccuTarget TM Negative Control sirna 는 non-targeting sirna 로써, human 과 mouse, rat 의알려진모든유전자들에대해 homology 가낮은 sequence 입니다. 따라서모든유전자의 knockdown 실험에 negative control 로사용할수있어서편리합니다. Transfection 효율을쉽게관찰하기위하여 sirna uptake 를확인할수있는 Fluorescent labeled Negative control sirna 도함께제공합니다 (Figure 8). 또한간편하고저렴한 positive 와 negative control sirna 로구성된 control set 도제공합니다. AccuTarget TM Positive Control sirna 와 Negative Control sirna 는바이오니아홈페이지에서원하는 scale 과 purification 을선택하여주문이가능합니다. Features and Benefits Excellent performance: 90% 이상의 knockdown 효율을갖는 positive control sirna Monitoring of transfection rate: 형광물질로 Labeling 되어편리하게효능검증이가능한 Negative control sets Competitive pricing: 경제적인비용으로극대화된효율을경험하세요. Positive control sirna 1. GAPDH-siRNA A) Northern blotting B) qrt-pcr Figure 5. Effects of Human GAPDH Positive Control sirna. HeLa cells were transfected separated with AccuTarget TM Human GAPDH Positive Control and Negative Control sirna using lipofectamine 2000 (Invitrogen) at a final concentration of 100 nm. Total cellular RNA was isolated from transfected cells 24 hr after transfection and subjected to Northern blot and Real-Time PCR analysis. As can be seen from Figure 5-A, about 3% GAPDH mrna remained. 2. GFP-siRNA BIONEER Figure 6. HeLa cells in a 24-well plate were co-transfected with 200 ng of CMV-GFP plasmid and 10 nm of GFP sirna using lipofectamine 2000 transfection reagent. Next day, the expression of GFP was observed by using a Nikon Eclipse TS100 epifluorescence microscope. In contrast to bright green fluorescence of GFP protein in NC-siRNA-transfected cells, no fluorescence was detected from GFP-siRNA-transfected cells, indicating efficient knockdown of GFP by using our positive control GFP-siRNA. 42

11 AccuTarget Control sirnas 3. Luciferase-siRNA Figure 7. HeLa cells in a 6-well plate were co-transfected with 400 ng of CMV-luc plasmid and 10 nm of luciferase sirna using lipofectamine 2000 transfection reagent. Next day, cells were harvested and assayed for luciferase activity. As shown in Figure. 7, co-transfection with our positive control luciferase sirna led to efficient knockdown of luciferase activity (85% - 95% knockdown compared to luciferase activity of NC-siRNAtransfected cells). 4. Mouse Positive control sirna Figure 8. Effects of Mouse Lamin A/C Positive Control sirna. Hepa1-6 cells were transfected separately with AccuTarget TM Mouse Lamin A/C Positive Control and Negative Control sirna using Lipofectamine RNAimax (Invitrogen) at a final concentration of 1 nm. Total cellular RNA was isolated from transfected cells 24 hr after transfection and subjected to Real-Time PCR analysis Figure 9. Effects of Mouse Ppib (Cyclophilin B) Positive Control sirna. Hepa1-6 & NIH 3T3 cells were transfected separately with AccuTarget TM Mouse Ppib (Cyclophilin B) Positive Control and Negative Control sirna using Lipofectamine RNAimax (Invitrogen) at a final concentration of 5 & 1 nm. Total cellular RNA was isolated from transfected cells 24 hr after transfection and subjected to Real-Time PCR analysis. 43

12 AccuTarget Control sirnas Negative control sirna Figure 10. HeLa cells transfected with FITC-labeled sirna (Cat. no. SN-1021) was observed by confocal microscopy. The fluorescent cells indicate that the target cells were successfully transfected with the sirna. Ordering Information AccuTarget TM Positive Control sirnas BIONEER Cat. no. Description Purification Guaranteed Yield Price ( 원 ) SP-1001 AccuTarget TM GAPDH Positive Control sirna Bio-RP 5 nmole 68,000 SP-1002 AccuTarget TM GAPDH Positive Control sirna Bio-RP 10 nmole 96,000 SP-1003 AccuTarget TM GAPDH Positive Control sirna Bio-RP 20 nmole 116,000 SP-1011 AccuTarget TM GAPDH Positive Control sirna HPLC 5 nmole 94,000 SP-1012 AccuTarget TM GAPDH Positive Control sirna HPLC 10 nmole 132,000 SP-1013 AccuTarget TM GAPDH Positive Control sirna HPLC 20 nmole 192,000 SP-2001 AccuTarget TM GFP Positive Control sirna Bio-RP 5 nmole 68,000 SP-2002 AccuTarget TM GFP Positive Control sirna Bio-RP 10 nmole 96,000 SP-2003 AccuTarget TM GFP Positive Control sirna Bio-RP 20 nmole 116,000 SP-2011 AccuTarget TM GFP Positive Control sirna HPLC 5 nmole 94,000 SP-2012 AccuTarget TM GFP Positive Control sirna HPLC 10 nmole 132,000 SP-2013 AccuTarget TM GFP Positive Control sirna HPLC 20 nmole 192,000 SP-3001 AccuTarget TM Luciferase Positive Control sirna Bio-RP 5 nmole 68,000 SP-3002 AccuTarget TM Luciferase Positive Control sirna Bio-RP 10 nmole 96,000 SP-3003 AccuTarget TM Luciferase Positive Control sirna Bio-RP 20 nmole 116,000 SP-3011 AccuTarget TM Luciferase Positive Control sirna HPLC 5 nmole 94,000 SP-3012 AccuTarget TM Luciferase Positive Control sirna HPLC 10 nmole 132,000 SP-3013 AccuTarget TM Luciferase Positive Control sirna HPLC 20 nmole 192,000 SP-4001 AccuTarget TM Mouse Lamin A/C Positive Control Bio-RP 5 nmole 68,000 SP-4002 AccuTarget TM Mouse Lamin A/C Positive Control Bio-RP 10 nmole 96,000 SP-4003 AccuTarget TM Mouse Lamin A/C Positive Control Bio-RP 20 nmole 116,000 SP-4011 AccuTarget TM Mouse Lamin A/C Positive Control HPLC 5 nmole 94,000 SP-4012 AccuTarget TM Mouse Lamin A/C Positive Control HPLC 10 nmole 132,000 SP-4013 AccuTarget TM Mouse Lamin A/C Positive Control HPLC 20 nmole 192,000 SP-5001 AccuTarget TM Mouse cyclophilin B c Positive Control Bio-RP 5 nmole 68,000 SP-5002 AccuTarget TM Mouse cyclophilin B c Positive Control Bio-RP 10 nmole 96,000 SP-5003 AccuTarget TM Mouse cyclophilin B c Positive Control Bio-RP 20 nmole 116,000 SP-5011 AccuTarget TM Mouse cyclophilin B c Positive Control HPLC 5 nmole 94,000 SP-5012 AccuTarget TM Mouse cyclophilin B c Positive Control HPLC 10 nmole 132,000 SP-5013 AccuTarget TM Mouse cyclophilin B c Positive Control HPLC 20 nmole 192,000 44

13 AccuTarget Control sirnas AccuTarget TM Negative Control sirnas Cat. no. Description Purification Guaranteed Yield Price ( 원 ) SN-1001 AccuTarget TM Negative Control sirna Bio-RP 5 nmole 68,000 SN-1002 AccuTarget TM Negative Control sirna Bio-RP 10 nmole 96,000 SN-1003 AccuTarget TM Negative Control sirna Bio-RP 20 nmole 116,000 SN-1011 AccuTarget TM Negative Control sirna HPLC 5 nmole 94,000 SN-1012 AccuTarget TM Negative Control sirna HPLC 10 nmole 132,000 SN-1013 AccuTarget TM Negative Control sirna HPLC 20 nmole 192,000 SN-1021 AccuTarget TM Fluorescein-labeled Negative Control sirna HPLC 5 nmole 134,000 SN-1022 AccuTarget TM Fluorescein-labeled Negative Control sirna HPLC 10 nmole 188,000 SN-1023 AccuTarget TM Fluorescein-labeled Negative Control sirna HPLC 20 nmole 264,000 AccuTarget TM Control sirna Sets Cat. no. Description Purification Guaranteed Yield Price ( 원 ) SS-1001 AccuTarget TM GAPDH Control sirna Set Bio-RP SS-1002 AccuTarget TM GFP Control sirna Set Bio-RP SS-1003 AccuTarget TM Luciferase Control sirna Set, Bio-RP SS-1011 AccuTarget TM GAPDH Control sirna Set HPLC SS-1012 AccuTarget TM GFP Control sirna Set HPLC SS-1013 AccuTarget TM Luciferase Control sirna Set HPLC 5 nmole positive control + 2 nmole negative control 5 nmole positive control + 2 nmole negative control 5 nmole positive control + 2 nmole negative control 5 nmole positive control + 2 nmole negative control 5 nmole positive control + 2 nmole negative control 5 nmole positive control + 2 nmole negative control 81,000 81,000 81, , , ,

14 AccuTarget Real-Time PCR Primer Library Description AccuTarget TM Human Validated Real-Time PCR Primer Library Set 는 Human genome 을대상으로 bioinformatics tool 을이용하여제작한후모든 primer 들의증폭효율을자사 Real-Time PCR 기기인 Exicycler TM 96 과 AccuPower GreenStar TM qpcr PreMix 를이용하여검증하였습니다. Real-Time PCR Primer Library Set 는유전자기능별 pathway 분류에따른 11,154 개의 primer set 로제공됩니다. SYBR Green (AccuPower GreenStar TM qpcr PreMix; Cat. no. K-6210) 을이용한 Real-Time PCR 에본제품을이용할경우더욱 specific 하고 sensitive 한결과를얻을수있습니다. (Figure 11) Features and Benefits In ready-to-ship format 11,154개 human 유전자특이적 primer All primers pre-validated: MALDI-TOF mass spectrometer를통한 QC검증 Competitive pricing: 경제적인비용으로극대화된효율을경험하세요. Figure 11. Real-Time PCR validation test of human Oxidoreductase using AccuTarget TM Human Oxidoreductase Real-Time PCR Primer set. Ordering Information AccuTarget TM Real-Time PCR Primer for Individual Gene Cat. no. Product Description Reactions Price ( 원 ) PHS-P01 AccuTarget TM Human Real-Time PCR Primer for Individual Gene 100 rxns 35,000 PHS-P02 AccuTarget TM Human Real-Time PCR Primer for Individual Gene 200 rxns 55,000 BIONEER 46

15 AccuTarget Real-Time PCR Primer Library AccuTarget TM Real-Time PCR Primer for Libraries Product Description no. of Genes Reactions / gene AccuTarget TM Human Antioxidant Real-Time PCR primer Set rxns AccuTarget TM Human Apoptosis Real-Time PCR primer Set rxns AccuTarget TM Human Cancer Real-Time PCR primer Set 1, rxns AccuTarget TM Human Caspase Real-Time PCR primer Set rxns AccuTarget TM Human Cell cycle Real-Time PCR primer Set rxns AccuTarget TM Human Cyclase Real-Time PCR primer Set rxns AccuTarget TM Human Cytochrome P450 Real-Time PCR primer Set rxns AccuTarget TM Human Deaminase Real-Time PCR primer Set rxns AccuTarget TM Human GPCR signaling pathway Real-Time PCR primer Set rxns AccuTarget TM Human Helicase Real-Time PCR primer Set rxns AccuTarget TM Human Isomerase Real-Time PCR primer Set rxns AccuTarget TM Human Kinase Real-Time PCR primer Set rxns AccuTarget TM Ligase Real-Time PCR primer Set rxns AccuTarget TM Human Kinase Real-Time PCR primer Set rxns AccuTarget TM Human Ligase Real-Time PCR primer Set rxns AccuTarget TM Human Lyase Real-Time PCR primer Set rxns AccuTarget TM Human Motor Real-Time PCR primer Set rxns AccuTarget TM Human NF-kB pathway Real-Time PCR primer Set rxns AccuTarget TM Human Nucleic acid binding Real-Time PCR primer Set 2, rxns AccuTarget TM Human Oxidoreductase Real-Time PCR primer Set rxns AccuTarget TM Human Peptidase Real-Time PCR primer Set rxns AccuTarget TM Human Phosphatase Real-Time PCR primer Set rxns AccuTarget TM Human Receptor Real-Time PCR primer Set 1, rxns AccuTarget TM Human Transporter Real-Time PCR primer Set rxns AccuTarget TM Human Tubulin Real-Time PCR primer Set rxns AccuTarget TM Human Ubiquitin Real-Time PCR primer Set rxns 47

16 sirna Oligonucleotide FAQs 1. 주문해서받게되는 sirna 의상태는어떤가요? Genome-wide predesigned sirna, validation sirna, sirna library, control sirna 는 sense 와 antisense 각각을동일농도로정제한다음 Annealing 과정을거쳐건조된상태의 duplex 로제공합니다. 이를고객이원하는 buffer 또는, 함께동봉해드리는 DEPC D.W. 를이용하여원하시는농도 (100 μm 로권장함 ) 로녹여서사용하시면됩니다. 6. Fluorescent dye modified sirna 는어떻게보관해야하나요? 빛에노출되면형광이약해질수있으므로빛이차단된용기에보관하시고또저장하는장소도빛이들어오지않는장소에서보관하시는것이좋습니다. Cy3, Cy5 는 ph 9 이상에서분해될수있으므로 Cy3, Cy5 modified 올리고는보관시염기성환경을피해야합니다. BIONEER Custom sirna 주문시 annealing service 를신청하시면 sense 와 antisense 각각을동일농도로정제한다음 Annealing 과정을거쳐 duplex 로제공해드립니다 ( 건조상태 ). 이를고객이원하는 buffer 또는, 함께동봉해드리는 DEPC D.W. 를이용하여원하시는농도 (50 μm 로권장함 ) 로녹여서사용하시면됩니다. Custom sirna 주문시 annealing service 를신청하지않으신경우 annealing buffer(1x) 를이용하여동봉되는 annealing protocol 대로 annealing 하시면됩니다. 2. In vitro 에서실험을하려고하는데어떤 scale 의 sirna 를주문하는것이좋을까요? (Purification 은어떤것을선택해야할까요?) 10 nmole scale 의 sirna 를 100 nm 로 transfection 한다면 6 well plate 기준으로 100 개 well 에 transfection 할수있는양입니다. In vivo 실험이아니라면 Bio-RP Purification 을선택하셔도좋은결과를얻으실수있습니다. In vivo 실험에서는 HPLC purification 을이용하시는것을권장합니다. 3. Genome-wide predesigned sirna 를주문하였는데효능이좋지않았습니다. 어떻게해야할까요? 먼저 sirna 를 PAGE 로확인해보시고 sirna 가분해되지않았다면 transfection 효율을확인해보셔야합니다. 한유전자당 3 개의 Predesigned sirna 를구매하신후그중 2 개가 80% 의 knockdown 효율을보이지않을경우, 2 개의 sirna 를무상으로제공해드립니다. 다만아래의 supported data 제출이필요합니다. 1. sirna 효율측정 data - NC : AccuTarget TM negative control, sirna 농도 : 100 nm 2. Transfection 효율측정 data - PC : AccuTarget TM GAPDH/GFP/Luciferase sirna - NC: AccuTarget TM Fluorescein labeled negative control 4. 합성된 sirna 에는 5 또는 3 위치에 phosphate 가붙어있나요? 주문하실때별도로선택하지않으면 5 과 3 위치에는 -OH 기가부착된상태로제공됩니다. 따라서 5 phosphate 가부착된 sirna 를원하시면 5 phosphorylation modification 된 sirna 를주문하셔야합니다. 5. sirna 는어떻게보관하며얼마나오래가나요? 일반적으로 sirna 는 -20 에서 1 년이상안정하다고볼수있습니다. 특히건조한상태에서는매우안정적이며오래보관하실수있습니다. 용액상태에서도안정하지만 RNase 오염등으로인해 degradation 될수도있습니다. 또한 sirna 는 얼렸다녹였다 를반복하게되면분해를촉진시켜품질이상이발생될수있으므로합성 sirna 는받으신후에필요량만큼여러개로분주하여각각을보관하신뒤하나씩꺼내어사용하시는것이바람직합니다. RNA 의경우, 높은 ph 에서 hydrolysis 에의해 phosphodiester bond 가깨질수있으니주의를요합니다. 7. 합성된 sirna 가몇 ng 인지알수있나요? 일반적으로 sirna 의합성은 guarantee nmole 단위로신청되며합성된결과도보통 nmole 단위로 report 됩니다. ng 또는 μg 단위로사용하시려면 mole 을 g 으로환산하시면됩니다. Report 에 sirna 의분자량이표시되기때문에쉽게바꿀수있습니다. 분자량 M.W. (g) X mole 수 (nmole) = sirna 의양 (ng) 8. sirna 실험을처음시작하려고합니다. 어떻게실험조건을잡을까요? sirna 실험에서중요한것은 sirna 가 cell 내에제대로 delivery 되는가하는점입니다. 현재판매되고있는 positive control 이나 NC-FITC 를이용하여간단하게 sirna delivery 를모니터링하실수있습니다. 9. sirna 실험을하기전에유념해야할점은무엇입니까? 첫째, sirna 염기서열이모두같은효율로유전자발현을억제하지않기때문에 3~4 개의 sirna 를시험하여가장적합한 sirna 를찾아야합니다. 둘째, 단백질수준에서억제효과가 sirna 작용에의한것임을정확히확인하기위해서 mrna 수준에서의측정도이루어져야합니다. 마지막으로원하는 knockdown phenotype 을확인하기위해한유전자당 2 개의다른 sequence 의 sirna 로실험하여같은 phenotype 이나타나는지확인해보는것이바람직합니다 nmole 의 sirna 로어떻게 100 nm 로 transfection 할수있을까요? 가장많이혼동하시는부분입니다. sirna 를 100 nm 로 6 well plate 의 1 well 에서 transfection 하실때, transfection volume 이 1 ml 이므로 100 nm(100 pmole/ml) 의농도로 sirna 를처리하려면 100 pmole 의 sirna 가필요합니다. 50 μm(50 pmole/μl) stock 의 sirna 를 2 μl 넣으면 100 pmole 의 sirna 를 transfection 한것이됩니다. 만약, 10 nmole 주문하시면 100 nm(100 pmole/ml) transfection 시 100 개 well 에사용하실수있습니다. 11. sirna 의 transfection efficiency 를확인하는방법은무엇인가요? NC-FITC 를 transfection 후형광현미경관찰을통해 transfection efficiency 확인이가능합니다. 또한 NC-FITC 는 sirna 와 transfection reagent 의사용농도및세포조건의확립등본실험에앞서실험조건을셋팅하기위한테스트시약으로사용될수있습니다. 12. sirna 의 knockdown 효율확인을위한실험방법은무엇인가요? qrt-pcr, Northern blot, Western blot 등의실험을통해 knockdown 효율을확인하실수있습니다 48

17 User Protocol Dilution Protocol 1. Briefly centrifuge tubes (or multi-well plates) containing sirna to ensure that the sirna pellet is located at the bottom of the tube. 2. Dissolve sirnas to a convenient stock concentration using the recommended volume of DEPC-D.W. (or RNase-free water) shown in Table Pipette the solution up and down 3~5 times (or vortex briefly). 4. Briefly centrifuge tubes (or multi-well plates) containing sirna to ensure that the solution is collected at the bottom of the tube. 5. Aliquot the sirnas into small volumes and store at -20. sirna is stable for 1 year under the specified storage condition. For best results, limit freeze-thaw events for each tube no more than five. Table 1. Recommended sirna resuspension volumes and concentrations. sirna Amount (nmol) DEPC-D.W. volume(μl) for desired final concentration 100 μm stock 20 μm stock Exceeds tube volume Transfection Protocol * We use the Lipofectamine TM RNAiMAX (Invitrogen; Cat.no ) and HeLa cell for transfection procedure. * This protocol is fixed at 6-well plate in vitro culture condition (if you want to change this condition, you have to consider the relative surface area (Table 2) and Invitrogen protocol, when you are seeding the cells into the culture dish). 1. One day (24 hr) before transfection, plate 3.0Ⅹ10 5 HeLa cells in each well with 2.5 ml of growth medium without antibiotics such that they will be 50~60% confluent at the time of transfection. 2. Remove the growth medium from the 6-well plate before transfection. And add the 500 μl fresh growth medium without serum in each well. 3. For each well to be transfected, prepare sirna duplex- Lipofectamine TM RNAiMAX complexes as follows Dilute sirna duplex (making final concentration as 5 nm~100 nm) in 250 μl growth medium (or Opti-MEM I Reduced Serum medium) without serum. Mix gently by vortex Mix Lipofectamine TM RNAiMAX gently before use, then dilute 3.5 μl in 250 μl medium (or Opti-MEM I Reduced Serum medium) without serum. Incubate this solution 5 minutes at room temperature Combine the diluted sirna duplex with the diluted Lipofectamine TM RNAiMAX. Mix and incubate for 20 minutes at room temperature. 4. Add the mixture to each well containing HeLa cells, which result 1 ml as total volume. Mix gently by hand rocking the plate back and forth. 5. Incubate the cells for 5~6 hr at 37 in CO2 incubator. 6. Change the medium with fresh one containing serum and incubate the cells 24~48 hr until you are ready to assay for gene knockdown. Table 2. The relative surface area of in vitro cell culture dish and culture media volume. Culture Vessel Relative surface area Volume of plating medium 96-well μl 48-well μl 24-well μl 6-well ml 60 mm 10 5 ml 100 mm ml 49

18 02 mirna AccuTarget TM mirnas 51 AccuTarget TM mirna mimics & inhibitors 52 AccuTarget TM mirna Controls 54

19 AccuTarget mirnas Overview MicroRNA(miRNA) 는 21~25 nucleotide(nt) 의 small non-coding RNA 분자로서진핵생물의유전자발현을제어하는조절물질입니다. 사람에는약 1700 여종의 mirna 가알려져있으며 (mirbase version 21), 이러한 mirna 는특정세포에서특정시점에발현되어 mrna 의발현을정교하게조절합니다. 주로 mrna 의 3 -UTR 에결합하여발생, 분화, 증식, apoptosis, stress response 등을포함한대부분의세포기능조절에관여하는것으로알려져있습니다. MicroRNA 의생성은두단계의과정으로이루어집니다. 최초의 microrna 전사체 (primary mirna/pri-mirna) 가핵안에서 Drosha 라는 RNase III type 효소에의해 70~90 nt 정도의 stemloop 구조의 pre-mirna 로만들어지고, 이후세포질로이동하여 Dicer 라는효소에의해절단되어 21~25 nt 의 mature mirna 로만들어집니다 (Figure 1). MicroRNA 는생명공학에새로운통찰력을제공하며, 비교적최근에발견된연구임에도불구하고, post-transcriptional level 에서가장중요한유전자 regulator 로인정되고있습니다. 여러연구에서 mirna 가 protein coding 유전자 30% 이상의발현을조절하는것으로밝혀지고있습니다. mirna 의 biogenesis, gene expression regulation mechanism 과기능에대한연구가축적되면서복잡한유전자조절네트워크를이해할수있는새로운지평이열렸다고인정되고있습니다. 최근연구를통해 mirna 는암발생단계나다양한암유형에따라고유한발현양상을보이고있으며많은질병및바이러스감염에중요한역할을할것으로알려졌습니다. 이러한결과는 mirna 가질병진단을위한새로운 biomarker 로서기능및 mirna 를이용한유전자치료에대한새로운전략을수행할수있다는것을보여줍니다. Figure 1. MicroRNA biogenesis (Annu. Rev. Cell Dev. Biol : ) 51

20 AccuTarget mirna mimics & inhibitors Description mirna 의안정성을극대화하기위해바이오니아의모든 mirna 는 clean room 에서제조됩니다. 또한 MALDI-TOF mass spectrometer 를통한엄격한 QC 과정에의해안정성이검증됩니다. 이렇게제작된고효율의 mirna 는고객의편의를위해주문다음날바로발송해드립니다. 바이오니아의 AccuTarget TM mirna mimic 은화학적으로합성되는 doublestranded RNA oligonucleotide 입니다. mirbase Sequence Database (version 21) 에존재하는 1700 여개의 Human Mature microrna 에대하여제조됩니다. AccuTarget TM mirna inhibitor 는 1700 여종의 human microrna 각각의 strand 를 targeting 할수있는 singlestranded synthetic inhibitor 입니다. 바이오니아는고객들의요구를충족시키기위해 96-well plate 에분주된다양한농도 (0.25, 0.5, 1, 2 nmole) 의 predesigned mimic 및 inhibitor 로구성된 mirna mimic, inhibitor library set 를제공합니다. 이러한 mirna mimic, inhibitor 는 5, 10 및 20 nmole guaranteed yield 로주문가능합니다. 또한, 고객이지정한염기서열을가지는 mimic 및 inhibitor 도제조가능하며이들로이루어진 flexible mirna library set 는최소 48 개이상의 mirna 로구매가능합니다. Ready-to-transfect mirna mimic 은 transfection 후세포에실제존재하는 endogenous mirna 와같은활성을나타내며, mirna inhibitor 는 target mirna 의활성을억제하여 mirna 의 loss-of-function 연구에사용될수있습니다. Figure 2. 바이오니아의 AccuTarget TM microrna mimics과타사제품의 reporter gene expression 제어비교평가. HEK cells을 96 well-plate 에 100 μl culture media 로배양후, transfection 실시. 50~70% confluence한상태에서 10 ng 의 luciferase vectors(3 UTR reporter와 control luciferase) 와 final concentration 100 nm의 microrna를 co-transfection 함. 24시간후, luciferase 활성을측정하여 translational repression을비교함. mtor와 Lin28b는각각 mir-100과 mir-125b의 validated target mrna. Features and Benefits Purification: 바이오니아의자동 HPLC 및 Bio-RP purification 방법으로높은품질의 mirna mimic 및 inhibitor 를보장합니다. Affordable pricing: 바이오니아는고품질의다양한 mirna 제품을경제적가격으로제공합니다. Synthesis and QC: 바이오니아의 mirna mimic 및 inhibitor 는완전자동화된 high-throughput mirna 생산시스템에의해 clean room 시설에서생산됩니다. 또한모든 mirna 제품은 MALDI- TOF Mass spectrometry analysis 에의해품질관리됩니다. 질량분석데이터는모든 mirna mimic 및 inhibitor 와함께제공됩니다. 또한 PAGE 분석을통해 mirna mimic 의 RNA 두가닥이제대로 annealing 되었는지확인합니다. 모든바이오니아의 mirna inhibitor 는 target mirna 의 complementary strand 로제공되며모든바이오니아의 mirna mimic 은 double-stranded RNA 로제공됩니다. 각각의 sense sirna 및 antisense sirna 는 MALDI-TOF analysis 에의해품질관리됩니다. annealing 된모든 doublestrand mirna 는 PAGE 분석을통해 annealing 을확인합니다. BIONEER 52

21 AccuTarget mirna mimics & inhibitors Ordering Information AccuTarget TM Custom mirnas Cat. no. Description Purification Guaranteed Yield Price ( 원 ) SMM-001 AccuTarget TM mirna mimic Bio-RP 5 nmole 98,000 SMM-002 AccuTarget TM mirna mimic Bio-RP 10 nmole 140,000 SMM-003 AccuTarget TM mirna mimic Bio-RP 20 nmole 165,000 SMI-001 AccuTarget TM mirna inhibitor Bio-RP 5 nmole 46,000 SMI-002 AccuTarget TM mirna inhibitor Bio-RP 10 nmole 68,000 SMI-003 AccuTarget TM mirna inhibitor Bio-RP 20 nmole 90,000 AccuTarget TM Library mirnas Cat. no. Description Purification Guaranteed Yield Price ( 원 ) SML-1001 AccuTarget TM mirna mimic Bio-RP 0.25 nmole 26,000 SML-1002 AccuTarget TM mirna mimic Bio-RP 0.5 nmole 33,000 SML-1003 AccuTarget TM mirna mimic Bio-RP 1 nmole 43,000 SML-1004 AccuTarget TM mirna mimic Bio-RP 2 nmole 55,000 SML-2001 AccuTarget TM mirna inhibitor Bio-RP 0.25 nmole 13,000 SML-2002 AccuTarget TM mirna inhibitor Bio-RP 0.5 nmole 17,000 SML-2003 AccuTarget TM mirna inhibitor Bio-RP 1 nmole 22,000 SML-2004 AccuTarget TM mirna inhibitor Bio-RP 2 nmole 28,000 단, Flexible library 주문의경우는 minimum 48 EA 이상주문시가능합니다 53

22 AccuTarget mirna Controls Description 바이오니아는 mirna mimic 기능연구의세포실험조건최적화를위해 AccuTarget TM mirna mimic Control 을제공합니다. mirna positive 및 negative Control 은바이오니아의 AccuTarget TM mirna mimic 을사용한 mirna 의기능연구실험에서대조군으로사용가능합니다. AccuTarget TM mirna Housekeeping Positive Control 은 housekeeping gene 인 GAPDH 의 3 UTR (untranslated region) 을 target 으로하며, transfection control 로사용할수있습니다. mirna mimic negative control sequence 는일반적인 mirna 의구조를가지며 human, mouse, rat 세포주를이용한실험에서 negative control 로사용될수있습니다. mirna Negative control 은모든 human, mouse, rat genomic sequence 및현재 mirbase Database 에있는 mirna sequence 에대해서서열유사성이최소화되어있음을 BLAST 를통해확인하였습니다. 바이오니아는두가지의 mirna mimic negative control 을제공합니다. 또한, positive 및 2 개의 negative mirna control 이포함된 AccuTarget TM mirna control set 가사용자의편의를위해제공됩니다. Features and Benefits Excellent performance: GAPDH 를 targeting 하며분명한 read-out 을 (90% 이상 knockdown efficiency) 제공하는 mirna housekeeping positive control 과 human, mouse, rat 에존재하는모든 mirna 에대해 minimum sequence identity 를갖는 mirna mimic negative control 을제공합니다. Purification: 바이오니아의자동 HPLC 및 Bio-RP purification 방법으로높은품질의 mirna mimic 및 inhibitor 를보장합니다. Affordable pricing: 바이오니아는고품질의다양한 mirna 제품을경제적인가격으로제공합니다. Figure 3. Performance of mirna mimics Positive and Negative controls. AccuTarget TM mirna Positive & Negative controls were transfected at 20 nm using Lipofectamine TM RNAiMAX into HeLa cell lines and assessed for their ability to decrease target mrna levels. Down-regulation of GAPDH was determined using the Real-Time quantitative RT-PCR at 48 hr post-transfection using Bioneer s Exicycler TM 96 qpcr instrument. BIONEER 54

23 AccuTarget mirna Controls Ordering Information AccuTarget TM Control mirnas Cat. no. Description Purification Guaranteed Yield Price ( 원 ) SMC-1001 AccuTarget TM mirna Housekeeping Positive Control (GAPDH) Bio-RP 5 nmole 68,000 SMC-1002 AccuTarget TM mirna Housekeeping Positive Control (GAPDH) Bio-RP 10 nmole 96,000 SMC-1003 AccuTarget TM mirna Housekeeping Positive Control (GAPDH) Bio-RP 20 nmole 116,000 SMC-2001 AccuTarget TM mirna mimic Negative Control #1 Bio-RP 5 nmole 68,000 SMC-2002 AccuTarget TM mirna mimic Negative Control #1 Bio-RP 10 nmole 96,000 SMC-2003 AccuTarget TM mirna mimic Negative Control #1 Bio-RP 20 nmole 116,000 SMC-3001 AccuTarget TM mirna mimic Negative Control #2 Bio-RP 5 nmole 68,000 SMC-3002 AccuTarget TM mirna mimic Negative Control #2 Bio-RP 10 nmole 96,000 SMC-3003 AccuTarget TM mirna mimic Negative Control #2 Bio-RP 20 nmole 116,000 SMC-4001 SMC-4002 SMC-4003 SMC-5001 SMC-5002 SMC-5003 AccuTarget TM Fluorescein-labeled mirna mimic Negative Control sirna #1 AccuTarget TM Fluorescein-labeled mirna mimic Negative Control sirna #1 AccuTarget TM Fluorescein-labeled mirna mimic Negative Control sirna #1 AccuTarget TM Fluorescein-labeled mirna mimic Negative Control sirna #2 AccuTarget TM Fluorescein-labeled mirna mimic Negative Control sirna #2 AccuTarget TM Fluorescein-labeled mirna mimic Negative Control sirna #2 HPLC 5 nmole 134,000 HPLC 10 nmole 188,000 HPLC 20 nmole 264,000 HPLC 5 nmole 134,000 HPLC 10 nmole 188,000 HPLC 20 nmole 264,000 SMC-2101 AccuTarget TM mirna inhibitor Negative Control #1 Bio-RP 5 nmole 68,000 SMC-2102 AccuTarget TM mirna inhibitor Negative Control #1 Bio-RP 10 nmole 96,000 SMC-2103 AccuTarget TM mirna inhibitor Negative Control #1 Bio-RP 20 nmole 116,000 SMC-3101 AccuTarget TM mirna inhibitor Negative Control #2 Bio-RP 5 nmole 68,000 SMC-3102 AccuTarget TM mirna inhibitor Negative Control #2 Bio-RP 10 nmole 96,000 SMC-3103 AccuTarget TM mirna inhibitor Negative Control #2 Bio-RP 20 nmole 116,000 SMC-4101 SMC-4102 SMC-4103 SMC-5101 SMC-5102 SMC-5103 AccuTarget TM Fluorescein-labeled mirna inhibitor Negative Control sirna #1 AccuTarget TM Fluorescein-labeled mirna inhibitor Negative Control sirna #1 AccuTarget TM Fluorescein-labeled mirna inhibitor Negative Control sirna #1 AccuTarget TM Fluorescein-labeled mirna inhibitor Negative Control sirna #2 AccuTarget TM Fluorescein-labeled mirna inhibitor Negative Control sirna #2 AccuTarget TM Fluorescein-labeled mirna inhibitor Negative Control sirna #2 HPLC 5 nmole 134,000 HPLC 10 nmole 188,000 HPLC 20 nmole 264,000 HPLC 5 nmole 134,000 HPLC 10 nmole 188,000 HPLC 20 nmole 264,

24 BIONEER 56