Characterization of swine leukocyte antigen polymorphism by sequence-based and PCR-SSP methods in Meishan pigs

Abstract

Resource herds of swine leukocyte antigen (SLA)-characterized pigs are an important tool for the study of immune responses, disease resistance, and production traits. They are also valuable large animal models for biomedical research, such as transplantation. The Meishan breed of pig is an economically significant breed that is available at several research institutions in the United States. We have characterized the SLA polymorphism of the breeding stock in the herd maintained at the University of Illinois and developed a simple assay to SLA type individuals within that herd. We have used a reverse transcription-polymerase chain reaction (RT-PCR)-based SLA typing method to clone and DNA sequence 19 SLA alleles at three SLA class Ia (SLA-1, SLA-2, and SLA-3) and two SLA class II (SLA-DRB1 and SLA-DQB1) loci. Based on this sequence information, a rapid SLA typing assay was developed to discriminate each allele using PCR with sequence-specific primers (PCR-SSP). Using this method, we were able to characterize the entire Meishan breeding stock and identify four SLA haplotypes present in the herd. The combination of SLA typing by cloning and DNA sequencing with PCR-SSP is therefore a valuable tool for the characterization of SLA alleles and haplotypes in resource herds of pigs.

Sequence variations of the locus‐specific 5′ untranslated regions of SLA class I genes and the development of a comprehensive genomic DNA‐based high‐resolution typing method for SLA‐2

“…Several methods have been used to characterize the polymorphisms of major SLA genes, such as cDNA cloning and sequencing (7)(8)(9), polymerase chain reaction with sequence-specific primers (PCR-SSP) (10)(11)(12), and genomic PCR and cloning (13). However, all these previously reported methods had inherent limitations, including the requirement of a DNA cloning step and incomprehensive allele identification.…”

Swine leukocyte antigen class II genes (SLA-DRA, SLA-DRB1, SLA-DQA, SLA-DQB1) polymorphism and genotyping in Guizhou minipigs

“…One novel class II haplotype was temporarily named as Hp-gz00.01 (SLA-DRA*gz01, SLA-DRB1*gz01, SLA-DQA*0101, SLA-DQB1*0601) and was confirmed in six pigs (G106, G108, G114, G338, G398, and G126). Another SLA class II haplotype that has been previously published and is present in Meishan pigs (Ho et al, 2006), and was officially named as Hp-00.14 (SLA-DRA*010103, SLA-DRB1*0901, SLA-DQA*0301, SLA-DQB1*0801), was confirmed in four pigs (G100, G118, G140, and G151). Crossover of class II haplotypes was observed in the G122 individual, and the crossover occurring between the Hp-gz0.14 and Hp-gz0.01 haplotypes, resulted in new recombinant haplotypes (SLA-DRA*gz01, SLA-DRB1*gz01, SLA-DQA*0101, SLA-DQB1*0601).…”

“…Each PCR-SSP reaction was multiplexed into an internal control (Ho et al, 2006), which amplifies a portion of the α-actin gene that is visible as a band at 498 bp. We found two SLA class II homozygote pigs (G423, G460, G465), which had the same SLA haplotype alleles (Hp-gz0.14, Hp-gz0.01) as their mothers (G100, G108) respectively.…”

Identification of swine influenza virus epitopes and analysis of multiple specificities expressed by cytotoxic T cell subsets

“…Experimental animals received chemically (C 3 H 4 O 2 ) inactivated swine influenza A virus of different strains given in equal volumes of Freund’s Incomplete adjuvant with 4 repeated immunizations at three-week intervals (Table 1). Initially, blood samples were collected from all pigs followed by SLA allele typing using PCR-SSP [14–16]. Candidate SwIV epitopes were selected using in silico predictions for binding by the online available NetMHCpan algorithm [17–19], and combined with previously mapped preferences expressed by SLA-1*0401 [13].…”