鸡传染性法氏囊病病毒BJQ902株感染性克隆的构建

doi:10.16431/j.cnki.1671-7236.2026.02.036

Abstract

Abstract:

Objective The purpose of this study was to construct infectious clone of Infectious bursal disease virus （IBDV） BJQ902 strain using reverse genetics technology， and provide important tools for vaccine development and the research of viral replication and pathogenic mechanisms. Method Based on the whole genome sequence of BJQ902 strain， PCR was used to amplify A and B fragments of IBDV BJQ902 strain’s genome. The RiboJ and HDVrz sequences were introduced at both ends of A and B fragments， respectively. Homologous recombination technology was employed to insert A and B fragments with ribozyme structures into vector pCAGGS， constructing the recombinant plasmids pCAGGS-BJQ-RAH and pCAGGS-BJQ-RBH. These vectors were co-transfected into DF-1 cells， and rBJQ902 strain was rescued after three consecutive passages in blind transmission. DF-1 cells were infected with both BJQ902 and rBJQ902 strains， and rBJQ902 strain was identified using PCR， Western blotting and indirect immunofluorescence assay （IFA）. Result The BJQ902 strain genome A and B fragments were successfully obtained， with sizes of 3 260 and 2 827 bp， respectively. The self-splicing ribozymes RiboJ and HDVrz sequences were ligated to the ends of fragment A and B， with sizes of 3 436 and 3 033 bp， respectively. The ribozyme-containing fragments A and B were then ligated to the pCAGGS vector， resulting in the successful acquisition of the recombinant plasmids pCAGGS-BJQ-RAH and pCAGGS-BJQ-RBH. rBJQ902 strain was obtained by blind passage for three consecutive generations in DF-1 cells. Compared with the parent strain BJQ902， rBJQ902 strain could also amplify the VP2 fragment after infecting DF-1 cells for 48 h. The lesions caused by rBJQ902 strain were consistent with those of the parent strain. Both Western blotting and IFA could detect the expression of VP2 protein in rBJQ902 strain， demonstrating that this strain exhibited similar biological characteristics to the parent virus during cell culture. Conclusion This study successfully obtained the infectious clone strain rBJQ902 of BJQ902 strain， providing technical support for in-depth research on the biological characteristics and pathogenic mechanism of IBDV and laying a foundation for subsequent virological research.

Key words: Infectious bursal disease virus （IBDV）; infectious clone; virus rescue; ribozyme

CLC Number:

S852.65⁺9.4

LYU Ran, YANG Hongwei, TU Min, MENG Zhaoying, JIN Huan, ZHANG Zhenhua. Construction of an Infectious Clone of Chicken Infectious Bursal Disease Virus Strain BJQ902[J]. China Animal Husbandry & Veterinary Medicine, 2026, 53(2): 916-925.

Figures/Tables 7

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References 28

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引物名称 Primer name	片段Segment	引物序列 Primer sequences （5'→3'）	退火温度 Annealing temperature/℃	用途 Purpose
IBDV-A-F	A	GGATACGATCGGTCTGACCC	55	全基因组扩增
IBDV-A-R	A	GGGGACCCGCGAACGGATCCA	55	全基因组扩增
pCAG-R-SeqA-F	A	GGCAAAGAATTCCTCGAGAGCTGTCACCGGATGTGCT	58	引入RiboJ核酶序列
pCAG-R-SeqA-R	A	GTTGGGTTTGATCTTGCAGGTTTGT	58	引入RiboJ核酶序列
SeqA-H-pCAG-F	A	AGGGCTGTCTCTGATGAGGACCTTG	58	引入HDVrz核酶序列
SeqA-H-pCAG-R	A	AGGAGTGAATTCCTCGAGTCGAGGCGGATCCGAGCTC	58	引入HDVrz核酶序列
IBDV-B-F	B	GGATACGATGGGTCTGACCCT	55	全基因组扩增
IBDV-B-R	B	GGGGGCCCCCGCAGGCGAAGG	55	全基因组扩增
pCAG-R-SeqB-F	B	GGCAAAGAATTCCTCGAGAGCTGTCACCGGATGTGCT	58	引入RiboJ核酶序列
pCAG-R-SeqB-R	B	CAGAGGGTCAGACCCATCGT	58	引入RiboJ核酶序列
SeqB-H-pCAG-F	B	CCCGGCCTTCGCCTGCGGGG	58	引入HDVrz核酶序列
SeqB-H-pCAG-R	B	AGGAGTGAATTCCTCGAGTCGAGGCGGATCCGAGCTC	58	引入HDVrz核酶序列
VP2-F	A	ATGACAAACCTGCAAGATCAA	55	鉴定VP2
VP2-R	A	CCTTATGGCCCGGATTATGTC	55	鉴定VP2

Construction of an Infectious Clone of Chicken Infectious Bursal Disease Virus Strain BJQ902

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