A novel strategy to attenuate porcine reproductive and respiratory syndrome virus by inhibiting viral replication in the target pulmonary alveolar macrophages via hematopoietic-specific miR-142

Table 1 Primers used for construction and identification of pWSK-JX-miR-142t

Primer^a	Sequence (5’-3’)^b	Position^c	Purpose
miR-142t-F	CCATAAAGTAGGAAACACTACAGAGGATCTGCTAAAACTAGCC	2493–2535	Amplification of miR142t
miR-142t-R	TGTAGTGTTTCCTACTTTATGGTCGAGCCCGCTCGGCAGCACG	2472–2514	Amplification of miR142t
JAF (SP6)	GCGGCCGCGCGATTTAGGTGACACTATAGATGACGTATAGGTGTTGGCTCTATG (NotI)	1–25	Fragment clone
JAR	GGCTTCGAAATTTGCCTGATCTT (BstBI)	4804–4826	Fragment clone
miR-142t	CCATAAAGTAGGAAACACTACA	2493–2514	Sequencing
W1F	ATGACGTATAGGTGTTGGCTCT	1–22	Sequencing
W1R	TACTCTTTCAGGAAGGGTGG	1556–1575	Sequencing
W2F	ACGCTCTGGTGCGACTACTA	1362–1381	Sequencing
W2R	AGGTTGTTCGGTTGTCTGATT	2233–2253	Sequencing
W3F	CCTCCGTGGCGCAACAAGTCTTG	2115–2137	Sequencing
W3R	CGATGATGGCTTGAGCTGAGTAT	3156–3178	Sequencing
W4F	TGAGCCTCTGGATTTGTCTGC	2952–2972	Sequencing
W4R	GGCGATCTCATTAGGAGCAGTT	4308–4329	Sequencing
W5F	TGCTTAGGCTTGGCATTGTTG	4214–4234	Sequencing
W5R	ACGGTGTTCAGTGAGGGCTTT	5544–5564	Sequencing

^aF denotes forward PCR primer; R denotes reverse transcription or reverses PCR primer
^bRestriction sites introduced by PCR are shown in boldface and specified in parentheses at the end of the sequence. And a viable chimeric virus designed as RvJX-miR-142t was successfully rescued
^cNumbers refer to nucleotide positions within the genome of JXwn06 (GenBank accession no: EF641008) as indicated. The underline denotes the miR142 target sequence (miR142t) at the first two lines and denotes the SP6 promoter followed by a nongenomic G (italicized) at the third line

ISSN: 2731-9970