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Table 2 PCR conditions for the detection and genotyping of avian respiratory viruses in samples from Pakistan

From: Molecular epidemiology of respiratory viruses in commercial chicken flocks in Pakistan from 2014 through to 2016

Virus

Primers

Sequences (5–3)

Target

Amplicon size (bp)

PCR conditions

Reference

Detection PCR

AAVV-1

FIP-1

5′ TACTTTGCTCACCCCCCTT 3’

Fusion gene (F)

280

94 C for 2 min; 40 cycles of 94 C for 30 s, 58 C for 30 s, 72 C for 1 min; final extension at 72 C for 5 min

[48]

FIP-2

5’ CATCTTCCCAACTGCCACT 3’

IBV

N791

5’ GTGATGACAAGATGAATGAGGA 3’

Nucleo-protein gene (N)

380

94 C for 2 min; 40 cycles of 94 C for 30 s, 54 C for 30 s, 72 C for 1 min; final extension at 72 C for 5 min

[49]

N1129

5’ CAGCTGAGGTCAATGCTTTATC 3’

ILTV

gEU

gEL

5’ GCTGGGTTCTGGGCTACACAAC 3′

5′ TGCGCGTGACTCGGAGAG 3’

Glyco-protein E gene (gE)

626

94 C for 2 min; 40 cycles of 94 C for 30 s, 61 C for 30 s, 72 C for 1 min; final extension at 72 C for 5 min

[50]

aMPV

G1a

5′ GGGACAAGTATCYMKAT 3’

Attachment glycoprotein gene (G)

441

94 C for 2 min; 40 cycles of 94 C for 30 s, 50 C for 30 s, 72 C for 1 min; final extension at 72 C for 5 min

[51]

G6a

5′ CTGACAAATTGGTCCTGATT 3’

AIV

M52C

5′ CTTCTAACCGAGGTCGAAAG 3’

Matrix gene (M)

280

95 C for 30 s; 40 cycles of 95 C for 30 s, 55 C for 30 s, 72 C for 30s; final extension at 72 C for 1 min

[52]

M253R

5’AGGGCATTTTGGACAAAKCGTCTA 3’

Genotyping PCR

AIV

HA-1134F

5′ GGAATGATHGAYGGNTGGTATG 3′

hemma-gglutinin gene (HA)

600

95 C for 30 s; 40 cycles of 95 C for 30 s, 55 C for 30 s, 72 C for 30s; final extension at 72 C for 1 min

[53]

NS-890 R

5′ ATATCGTCTCGTATTAGTAGAAAC AAGG 3′

IBV

S15

5′ TGAAAACTGAACAAAAGACA 3’

Spike gene (S)

700

95 °C for 2 min; 40 cycles of 95 °C for 30 s, 52 °C for 30 s, 72 °C for 30 s; final extension of 72 °C for 12 min

[54]

CK2

5′ CTCGAATTCCNGTRTTRTAYTGRCA 3′

  1. bp base pairs
  2. athis primer pair does not detect serotype C viruses