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Building a Custom RT2 Profiler PCR Array in GeneGlobeSamuel RulliGlobal Product Manager – RT2 Profiler PCR Arrays Samuel.rulli@qiagen.com

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How to build a custom array for RNA-seq validation

Gene Transcript Fold Change p-val q-valIL12B NM_002187 Infinity 4.97E-04 1.20E-03TNFSF14 NM_172014 129 8.99E-55 1.49E-52CXCL2 NM_002089 110 3.26E-30 1.08E-28IL1RN NM_173843 102 7.30E-51 6.06E-49C3 NM_000064 82.4 1.66E-18 2.13E-17CXCL9 NM_002416 47.1 5.51E-39 3.05E-37FASLG NM_000639 35.1 2.33E-22 3.86E-21CCL11 NM_002986 35 4.86E-11 2.99E-10SELE NM_000450 26.5 1.22E-27 2.52E-26TNFSF10 NM_003810 23.3 2.24E-29 5.31E-28CD27 NM_001242 19.5 1.09E-17 1.21E-16CCL5 NM_002985 18.6 2.53E-27 4.67E-26SELP NM_003005 17.3 6.77E-18 8.02E-17CD80 NM_005191 12.7 2.28E-17 2.36E-16CASP8 NM_033355 11.3 2.76E-17 2.69E-16IFNG NM_000619 10.8 8.81E-07 3.18E-06IL1R1 NM_000877 9.9 1.64E-16 1.43E-15CXCL10 NM_001565 9 4.80E-15 3.46E-14BCL3 NM_005178 8.9 1.33E-19 1.84E-18CCL22 NM_002990 8 3.69E-04 9.01E-04CCNB2 NM_004701 7.7 2.10E-12 1.40E-11MYD88 NM_002468 7.7 3.00E-17 2.77E-16IL2 NM_000586 7.3 3.24E-03 6.47E-03HMOX1 NM_002133 6.8 2.86E-15 2.16E-14TNFRSF10B NM_147187 6.5 1.93E-11 1.23E-10LTBR NM_002342 6.2 1.02E-13 7.09E-13

VCAM1 NM_001199834 5.6 2.66E-10 1.52E-09CD69 NM_001781 5.3 1.88E-07 7.24E-07TLR9 NM_017442 -5.00 4.57E-08 1.95E-07CD83 NM_004233 -13.33 1.61E-21 2.44E-20CCNE1 NM_001238 -14.93 5.97E-16 4.96E-15CCNG2 NM_004354 -15.15 7.87E-31 3.27E-29CCND2 NM_001759 -17.54 8.22E-30 2.27E-28IFNA1 NM_024013 -128.21 2.76E-15 2.16E-14

52 samples with 48 genes

34 genes +/- 5 fold change

5 normalization genes

Controls for RT, PCR and gDNA contamination

n=1 qPCR assays per sample

Biorad CFX96 instrument

26 total plates

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How to Find the Custom RT2 PCR Array builder

1.) www.qiagen.com/MyPCRArray (capitalization not important)

2.) Navigate from QIAGEN.com homepage

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Genes to upload

IL12B,TNFSF14,CXCL2,IL1RN,C3,CXCL9,FASLG,CCL11,SELE,TNFSF10,CD27,CCL5,SELP,CD80,CASP8,IFNG,IL1R1,CXCL10,BCL3,CCL22,CCNB2,MYD88,IL2,HMOX1,TNFRSF10B,LTBR,VCAM1,CD69,TLR9,CD83,CCNE1,CCNG2,CCND2,IFNA1

AIRN,BANCR,FAS-AS1,PCAT1,TUG1,XIST

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Type in NCBI GeneSymbol, Entrez GeneIDs, NCBI Transcript Refseq IDs. SangerIDs, Sanger Accessions GeneGlobe Catalog Numbers

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Confirm gene list

Confirm gene list

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Configure Product

Select instrument

Choose plate

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Add controls and normalization genes

! Select RT2 for only mRNA applications. Select RT2 lncRNA for lncRNA applications. Select both to have option of mRNA or lncRNA from same gene locus.

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Configure Plate (add, move or delete genes)

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I added some controls and normalization genes

Price is for minimal # platesAdd to cart to assign Custom catalog number

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How many plates do I need? 15 (and I can order exactly 15!)

Material # 330171 Material # 330181

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My shopping cart. Order online or call customer care.

To order online: press“Checkout” and follow ordering process (credit card or PO)

To order using customer care, call and mention product 330171. Quantity 1. CLAH20268.12 plates

Product 330181. Quantity 3. CLAH20268

Forward cart to procurement

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Ordering process for 330171/330181

Ordering online:

Customer or procurement can use credit card or PO. Once order is placed it automatically enters SAP and once order has cleared SAP it enters production.

Calling customer care:

Customer must supply custom array number and total number of plates.

Custom Array numbers

Will start with CLA[SPECIES] ##### (5 unique numbers).

One gene list to one unique catalog number

Any changes to gene list generates new unique catalog number

Customer’s orders and gene lists will be saved online for future re-orders