OVERCOMING mRNA PRODUCTION SCALE-UP CHALLENGES mRNA 3' Poly(A) characterization Accurate, sensitive detection of poly(A) distribution cap UTR 5' CDS UTR Poly(A) tail AAAA 3' * Protects against exonuclease degradation. Poly(A) tails (117-134 As) * Promotes translation. Nucleic acid candidate RNase T1 Oligonucleotides Poly(A) tail 123A Oligo d(T) 25 cleanup Poly(A) tail 100 20 40 60 80 3 4 Time (min) Poly(A) tails are baseline separated from the rest of mRNA digest. 5 6 38500 39000 39500 40000 40500 41000 41500 Mass Poly(A) species with different lengths are characterized with high sensitivity and high mass accuracy (mass error <20 ppm) in the deconvoluted spectrum. Their relative abundance can be estimated using peak height. 42000 42500 43000 43500 44000 100 122A 10 20 30 40 50 60 70 80 90 121A 127A 124A 125A 126A View complete application note Poly(A) tail features * Length is transcript dependent (100-250 nucleotide long). 120A 128A 129A 119A 118A 117A 130A 131A 132A 133A 134A Bf Thermo Scientific™ DNAPac™ RP HPLC Columns Vanquish Flex 300 32.25 H x 21.65 W Thermo Scientific™ Vanquish™ Flex UHPLC System Acclaim C18 Column Columns 35 Thermo Scientific™ Orbitrap Exploris™ 240 Mass Spectrometer Orbitrap Exploris 240 Thermo Scientific™ BioPharma Finder Software Relative abundance Relative intensityhttps://www.thermofisher.com/us/en/home.html https://assets.thermofisher.com/TFS-Assets/CMD/Application-Notes/an-001183-bt-orbitrap-exploris-240-ms-mrna-hram-lc-ms-an001183-na-en.pdf https://www.thermofisher.com/order/catalog/product/IQLAAAGABHFAPUMBJC?SID=srch-srp-IQLAAAGABHFAPUMBJC https://www.thermofisher.com/order/catalog/product/088923 https://www.thermofisher.com/order/catalog/product/BRE725535?SID=srch-srp-BRE725535 https://www.thermofisher.com/order/catalog/product/OPTON-30986?SID=srch-srp-OPTON-30986