Evaluation of custom multiplex real - time RT - PCR in comparison to fast - track diagnostics respiratory 21 pathogens kit for detection of multiple respiratory viruses

Malhotra, Bharti; Swamy, M. Anjaneya; Reddy, P. V. Janardhan; Kumar, Neeraj; Tiwari, Jitendra Kumar

doi:10.1186/s12985-016-0549-8

Virology Journal

Table 1 Custom primers and probes used for the detection of respiratory viruses

From: Evaluation of custom multiplex real - time RT - PCR in comparison to fast - track diagnostics respiratory 21 pathogens kit for detection of multiple respiratory viruses

VIRUS	Froward primer (5’ - 3’)	Reverse primer (5’ - 3’)	Probe (5’ - 3’)^d	References
Panel-1
Flu B	GAGACACAATTGCCTACCTGCTT	TTCTTTCCCACCGAACCAAC	^aFAM – AGAAGATGGAGAAGGCA AAG CAGA ACTAGC	Esposito et al., 2010 [17]
HCoV 229E	CAGTCAAATGGGCTGATGCA	AAAGGGCTATAAAGAGAATAAGGTATTCT	^bVIC – CCCTGACGACCACGTTGTGGTTCA	Hammit et al., 2011 [6]
HCoV OC43	CGATGAGGCTATTCCGACTAGGT	CCTTCCTGAGCCTTCAATATAGTAACC	^cNED-TCCGCCTGGCACGGTACTCCCT	Hammit et al., 2011 [6]
Panel-2
HPIV-4	CAGAYAACATCAATCGCCTTACAAA	TGTACCTATGACTGCCCCAAARA	^aFAM – CCMATCACAAGCTCAGAAATYCAAAGTCGT	Hammit et al., 2011 [6]
HPIV-1	GTGATTTAAACCCGGTAATTTCTCA	CCTTGTTCCTGCAGCTATTACAGA	^bVIC- ACCTATGACATCAACGAC	Hammit et al., 2011 [6]
HPIV-3	CCAGGGATATAYTAYAAAGGCAAAA	CCGGGRCACCCAGTTGTG	^cNED – TGGRTGTTCAAGACCTCCATAYCCGAGAAA	Hammit et al., 2011 [6]
Panel-3
Influenza A(H1N1) pdm09	GTGCTATAAACACCAGCCTYCCA	CGGGATATTCCTTAATCCTGTRGC	^aFAM - CAGAATATACATCCRGTCACAATTGGARAA	WHO, 2009 [22]
HRV	TGGACAGGGTGTGAAGAGC	CAAAGTAGTCGGTCCCATCC	^bVIC - TCCTCCGGCCCCTGAATG	Hammit et al., 2011 [6]
HPIV-2	ATGAAAACCATTTACCTAAGTGATGGA	CCTCCYGGTATRGCAGTGACTGAAC	^cNED - TCAATCGCAAAAGC	Hammit et al., 2011 [6]
Panel-4
RSV A/B	GGAAACATACGTGAACAAGCTTCA	RSV-A: CATCGTCTTTTTCTAAGACATTGTATTGA	^aFAM – TGTGTATGTGGAGCCTT	Kwofie et al., 2012 [10]
RSV A/B	GGAAACATACGTGAACAAGCTTCA	RSV-B: TCATCATCTTTTTCTAGAACATTGTACTGA	^aFAM – TGTGTATGTGGAGCCTT	Kwofie et al., 2012 [10]
HCoV NL63	ACGTACTTCTATTATGAAGCATGATATTAA	AGCAGATCTAATGTTATACTTAAAACTACG	^bVIC –ATTGCCAAGGCTCCTAAACGTACAGGTGTT	Hammit et al., 2011 [6]
HCoV HKU1	AGTTCCCATTGCTTTCGGAGTA	CCGGCTGTGTCTATACCAATATCC	^cNED - CCCCTTCTGAAGCAA	Cui et al., 2011 [11]
Panel-5
EV	CCCTGAATGCGGCTAATCC	ATTGTCACCATAAGCAGCCA	^aFAM- AACCGACTACTTTGGGTGTCCGTGTTTC	Wolffs et al., 2011 [23]
HPeV	GTAACASWWGCCTCTGGGSCCAAAAG	GGCCCCWGRTCAGATCCAYAGT	^bVIC- CCTRYGGGTACCTYCWGGGCATCCTTC	Nix et al., 2008 [12]
HBoV	TGCAGACAACGCYTAGTTGTTT	CTGTCCCGCCCAAGATACA	^cNED – CCAGGATTGGGTGGAACCTGCAAA	Sanghvi et al., 2012 [22]
Panel-6
Flu A	GACCRATCCTGTCACCTCTGAC	AGGGCATTYTGGACAAAKCGTCTA	^aFAM – TGCAGTCCTCGCTCACTGGGCACG	WHO, 2009 [22]
HAdV	GCCCCAGTGGTCTTACATGCACATC	GCCACGGTGGGGTTTCTAAACTT	^bVIC – TGCACCAGACCCGGGCTCAGGTACTCCGA	Hammit et al., 2011 [6]
HMPV A/B	CATCAGGTAATATCCCACAAAATCAG	GTGAATATTAAGGCACCTACACATAATAARA	^cNED - TCAGCACCAGACACAC	Sanghvi et al., 2012 [13]

NOTE: The lower limit for the detection of HBoV- 1 DNA copy/ml, HMPV- 30 Rna copies/ml, HPeV- 10³ (cell culture infective dose) CCID₅₀ - 10⁴ CCID₅₀, RSV A/B- 2×10⁴copies/μl, HCoV HKU1- 5×10³ copies/ml, Flu B- 2.2 Log₁₀ (viral particles) VP/ml, and Influenza A(H1N1)pdm09 - 2×10¹ to 2×copies/ml
^aFAM - Detection wavelength - 518 nm; ^bVIC detection wavelength - 554 nm; ^cNED detection wavelength 575 nm
^dAll the probes were having (non fluorescence quencher) NFQ as quencher at 3’ end

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ISSN: 1743-422X

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