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Abstract
All xenografts from pigs impose infection risk by porcine endogenous retrovirus (PERV). The purpose of this study was to investigate the env constructs with the comparison of the ratio of the competent form to the defective one of env in subtypes, PERV-A, PERV-B and PERV-C in different pig breeds. The results of PCR amplification of env represented that all env subtypes had more than two defective forms which cannot bind to host cells due to the absence of binding regions of env in miniature pigs, SNU and PWG, and farm pig breeds, Duroc, Yorkshire and Landrace. In addition, comparing the full sequences with the defective ones in three subtypes demonstrated that the present percentages of env sequences in defective PERV-A, PERV-B and PERV-C were approximately 50%, 38∼45% and 4∼11%, respectively, in SNU and PWG pigs whereas PERV-A and PERV-B occupied around 40 to 60% but PERV-C was not detected in farm pigs. Quantitative real-time PCR assays with primers and probes targeted to proline-rich region (PRR) of each env subtype were done to measure the copy numbers of each env subtype. When the reference was set with copy number of PERV-A, the ratio of those of PERV-B and PERV-C to the reference were 1.5 to 6.0 folds high in SNU and PWG pigs while 1.0 or less in farm pigs. These contradictory results of PERV-C constructs and copy numbers in SNU pigs suggests that many truncated or short defective sequences of PERV-C might be present in them.
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Figure 1.
Amplified profile of env subtypes in miniature pigs (SNU and PWG) and farm pig breeds (Duroc, Yorkshire and Landrace). Gene amplification was done by PCR with the primers described in Table 1. All the experiments revealed the similar amplification patterns and the representative pictures were shown. (M) denotes molecular marker; (A), (B) and (C) denote PERV subtypes A, B and C, respectively.
Figure 2.
Schematic diagrams of constructs of PERV A, B and C env gene derived from SNU miniature pig. SU, surface; TM, transmembrane; RBD, receptor binding domain; VRA, variable region a; VRB, variable region b; PRR, proline-rich region; open triangle: stop codon.
Figure 3.
Pylogenetic relationship tree using neighbor-joining methods on the nucleotide sequences of complete and defective PERV env derived from SNU pigs.
Figure 4.
The proportion of the full length sequences and defective ones of env in 18 SNU and 11 PWG miniature pigs. The relative amount of each construct was measured by the densitometry of amplified PCR products on agarose gel. Results were shown with mean + standard deviation.
Figure 6.
The proportion of the env A, B and C subtypes in various pig breeds. (A) the percentages of each env subtype by densitometry of PCR product on the agarose gel. (B) the ratio of copy numbers of PERV-B and PERV-C to that of PERV-A. Results were shown in mean + standard deviation in 18 SNU, 11 PWG, 3 Duroc, 3 Yorkshire, and 3 Landrace pigs.
Table 1.
PERV env specific primers and probes used in this study.
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