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Primers and probes used for rAAV quantitative analysis

ApplicabilityTargetSequences
Universal
All rAAV samples
ITR2primers: 5′-ggaacccctagtgatggagtt-3′ and
5′-cggcctcagtgagcga-3′;
probe: R-5′-cactccctctctgcgcgctcg-3′-Q.
Semi-universal
Group of rAAV samples
CMV promoterprimers: 5′-ttcctacttggcagtacatctacg-3′ and
5′-gtcaatggggtggagacttgg-3′;
probe: R-5′-tgagtcaaaccgctatccacgccca-3′-Q;
and other sequences.
CAG promoterprimers: 5′-ctgaccgcgttaatcccaca-3′ and
5′-acaagccgtgattaaaccaaga-3′.
CBA promoterprimers: 5′-ccgcagccattgccttt-3′ and
5′-ccgcacagatttgggacaa-3′;
probe: R-5′-atggtaatcgtgcgagagggcgc-3′-Q.
TBG promoterprimers: 5′-aaactgccaattccactgctg-3′ and
5′-ccataggcaaaagcaccaaga-3′;
probe: R-5′-ttggcccaatagtgagaactttttcctgc-3′-Q.
GRK1 promoterprimers: 5′-tctcttaaggtagccccgg-3′ and
5′-atccgattagatcattctgccc-3′;
probe: R-5′-cctcacttttcccctgagaaggaca-3′-Q.
RBG intronprimers: 5′-tcaggtgcaggctgcctat-3′ and
5′-tttgtgagccagggcattg-3′;
probe: R-5′-agaaggtggtggctggtgtgg-3′-Q.
CMV enhancerprimers: 5′-gtcaatgggtggagtatttacgg-3′ and
5′-gcattatgcccagtacatgacct-3′;
probe: R-5′-caagtgtatcatatgccaagtacgccccc-3′-Q;
and other sequences.
WPREprimers: 5′-ttggatgctcgcctgggttg-3′ and
5′-aggaaggtccgctggatcga-3′.
SV40 polyAprimers: 5′-agcaatagcatcacaaatttcacaa-3′ and
5′-ccagacatgataagatacattgatgagtt-3′;
probe: R-5′-agcatttttttcactgcattctagttgtggtttgtc-3′-Q.
BGH polyAprimers: 5′-catataaaatgaggaaattgcatcgca-3′ and
5′-tcagaacccatagagcccaccg-3′;
and other sequences.
RBG polyAprimers: 5′-gatttttcctcctctcctgactactc-3′ and
5′-gctgcaggtcgagggatct-3′;
probe: R-5′-cagtcatagctgtccctcttctctt-3′-Q;
and other sequences.
Individual
Unique rAAV samples
eGFPprimers: 5′-cacccacgtgaccacccttac-3′ and
5′-ggatgttgcagtcctccctg-3′;
and other sequences.
hrGFPprimers: 5′-gatccgcagcgacatcaacc-3′ and
5′-gtacaccacctcgaagctgg-3′;
probe: R-5′-gaggagatgttcgtgtaccgcgtgg-3′-Q.
emGFPprimers: 5′-acggcgacgtaaacggccac-3′ and
5′-gcgaagcactgcacgccgta-3′.
pU6primers: 5′-gggaaataggccctcttcctgccc-3′ and
5′-caccacgtgacggagcgtgac-3′.
ITR5primers: 5′-cccccccaaacgagccag-3′ and
5′-acccccttgcttgagag-3′;
probe: R-5′- cgagcgaacgcgacaggggggagagtg-3′-Q.

R—probe reporter, Q—probe quencher, BGH—bovine growth hormone, CAG—cytomegalovirus enhancer/chicken β-actin, CBA—chicken β-actin, CMV—Cytomegalovirus, eGFP—enhanced green fluorescent protein, emGFP—emerald green fluorescent protein, GRK1—rhodopsin kinase, hrGFP—humanized Renilla reniformis green fluorescent protein, ITR—inverted terminal repeat, polyA—polyadenylation signal, pU6—U6 promoter, RBG—rabbit β-globin, SV40—Simian virus 40, TBG—human thyroxine-binding globulin and WPRE—woodchuck hepatitis B virus posttranscriptional regulator

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