Characterization of a novel allelic variant in HLA‐B*40 lineage,HLA‐B*40:298:02, by cloning and sequencing

A novel allelic variant in HLA‐B*40 lineage, HLA‐B*40:298:02, has been identified in an individual of Han ethnicity afflicted with nasopharyngeal carcinoma in Hunan province, southern China. Following polymerase chain reaction–Sanger sequence‐based typing (PCR–SBT), this new variant was further confirmed by two distinct strategies of cloning and sequencing. HLA‐B*40:298:02 differs from HLA‐B*40:298:01 by a single synonymous cytosine substitution at nucleotide position 26 (T→C) in exon 3, which corresponds to codon 99 of the mature HLA‐B mRNA molecule. This new allele has an estimated frequency of 0.0002, in about 2,500 sequence‐based typed subjects from the same population.     

Characterization of a new HLA‐A allele,HLA‐A*02:07:08, by cloning and sequencing

In this report, we present a novel HLA‐A*02:07 allele, HLA‐A*02:07:08. HLA‐A*02:07:08 was identified in an individual of Han ethnicity in Hunan province, southern China. Following polymerase chain reaction‐sequence‐based typing (PCR‐SBT), this new allele was further confirmed by cloning and sequencing. HLA‐A*02:07:08 differs from HLA‐A*02:07:01 by a single synonymous C to T substitution at nucleotide position 131 in exon 3.     

Characterization of a novel HLA‐B*39:01:01‐related allele,HLA‐B*39:130, by cloning and phasing

A novel HLA‐B*39:01:01‐related variant, HLA‐B*39:130, has been identified in a normal individual of Han ethnicity in Hunan province, southern China. Following Sanger polymerase chain reaction–sequence‐based typing (PCR‐SBT), this new allele was further confirmed by cloning, phasing and sequencing. Aligned with HLA‐B*39:01:01, HLA‐B*39:130 has a nonsynonymous thymine substitution at nucleotide position 94 in exon 4, resulting in amino acid change from threonine to isoleucine at codon 214 (ACA→ATA) of the mature HLA‐BmRNA molecule.     

A novel HLA‐B*40 allele,B*40:01:40, identified in a Chinese individual

A new allele, officially named B*40:01:40, was detected in a Chinese individual by sequence‐based typing (SBT). The new allele differs from B*40:01:01 by a single nucleotide exchange at position 99 in codon 9, which results in synonymous substitution and seems not to compromise the HLA complex and T‐cell receptor interaction.     

Identification and characterization of a novel HLA‐A allele,HLA‐A*68:105, by genomic sequencing

A novel HLA‐A allele, HLA‐A*68:105, was detected by sequence‐based typing (SBT) in an Italian bone marrow donor. It differs from HLA‐A*68:01:02 at five nucleotides, three intronic, nt 699 T‐>G (intron 2), nt 705 T‐>C (intron 2) and nt 2770 G‐>A (intron 7), and two located in exon 3, at positions 726 A‐G (codon 94 Ile‐>Val) and 733 T‐G (codon 97 Arg‐>Met), respectively.     

Characterization of the novel HLA‐DQB1*06:48 allele by group‐specific sequencing

HLA‐DQB1*06:48 has single nucleotide polymorphisms within codons 70 and 62 of exon 2 (GGG>AGG and AAG>AAC) relative to HLA‐DQB1*06:02:01 and HLA‐DQB1*06:37. This results in amino acid differences (G>R and K>N) that will change the polarity and charge of the encoded antigen and may therefore affect its peptide repertoire.     

Characterization of a novel MICA allele,MICA*012:05, by cloning and sequencing

A new MICA allelic variant, MICA*012:05, has been identified in a Chinese Mongolian population. Following polymerase chain reaction–sequence‐based typing (PCR‐SBT), this new allele was further confirmed by cloning and sequencing. MICA*012:05 was linked to an HLA‐A*24‐C*01‐B*55:02‐DRB1*09 haplotype. MICA*012:05 differs from MICA*012:01 by a single synonymous C to T substitution at nucleotide position 269 in exon 3.     

Identification of a new HLA‐G allele,HLA‐G*01:19, by cloning and phasing

A new HLA‐G allelic variant, HLA‐G*01:19, was identified in a southern Chinese Han population by polymerase chain reaction–sequence‐based typing (PCR‐SBT), cloning and phasing. HLA‐G*01:19 differs from HLA‐G*01:04:01 by a nonsynonymous cytosine at position 99 in exon 2, resulting in amino acid change from valine to leucine at codon 34 of the mature HLA‐G molecule.     

A novel HLA‐B allele,HLA‐B*35:279, identified by sequencing‐based typing in a Czech patient

The identification of a novel HLA‐B*35:279 allele in a Czech patient is described. This allele is identical to the B*35:03:01 variant except the G/A nucleotide exchange at position 652 of the HLA‐B gene that corresponds to the amino acid substitution from valine to isoleucine in alpha 3 domain of the HLA‐B antigen.     

Characterization of three new HLA Class I Alleles in Spanish Caucasians,HLA‐A*02:620, HLA‐B*27:150 and HLA‐B*07:05:01:02

Three new HLA class I alleles, HLA‐A*02:620, HLA‐B*27:150 and HLA‐B*07:05:01:02, were described in the Spanish Caucasoid population.     

Immunogenetics of a new HLA‐B null allele,HLA‐B*4423N

The second example of an HLA‐B*44 null allele (B*4423N) was identified by discrepancies between serological and polymerase chain reaction–sequence‐specific primer (PCR‐SSP) typing in two north‐western European Caucasoid unrelated stem cell donor volunteers. HLA‐B*4423N was identical to B*440201 except for a single nucleotide substitution at position 493 in exon 3, resulting in a premature stop codon at bases 493–495 (TAG rather than CAG at codon 141). As expected, comprehensive serological testing using 54 antisera, directed towards B44 or Bw4, failed to identify the HLA‐B44 (Bw4) specificity. The B*4423N‐bearing haplotype was identified as A*0201, Cw*0501, DRB1*0408, DRB4*01, DQA1*03, DQB1*0304 and the frequency of B*4423N estimated as 0.00006 (carriage frequency 0.0121%) in 16 533 subjects resident in Wales.     

Two novel HLA‐C alleles,HLA‐C*02:02:34 and HLA‐C*03:369, were identified in the same individual

Two new HLA class I alleles, HLA‐C*02:02:34 and HLA‐C*03:369, were characterized in a single Polish bone marrow donor.     

Identification and characterization a novel HLA‐A allele,A*02:355, by sequence‐based typing in a Chinese potential donor

The new allele A*02:355 differs from A* 02:03:01 at positions 98 (T→A) and 102(A→C) resulting in an amino acid exchange F9→T. Interallelic sequence exchange is more likely the mechanism of its origination. The amino acid replacement influences the HLA peptide binding cleft and might have significant functional effects.     

A novel HLA-A*02 allele, A*02202, identified by nucleotide sequencing

In this paper we report the identification of a new HLA-A*02 allele in two members of an Afghan family. This novel allele, designed as A*02202, differs from A*02201 by a silent substitution at codon 66 (AAC-->AAT) in the alpha1 domain. A*02202 appears to be the result of a novel mutation (Note).     

Identification of a novel MICB allele,MICB*030, by cloning and sequencing

A novel MICB allele, MICB*030, has been identified in a healthy Chinese individual of Mongol ethnicity residing in northern China by polymerase chain reaction sequence‐based typing (PCR‐SBT) and confirmed by cloning and sequencing. MICB*030 was linked to HLA‐B*35. Aligned with MICB*005:02, MICB*030 has a nonsynonymous adenine substitution at nucleotide position 50 in exon 3, leading to amino acid change from serine to arginine at codon 102 of the mature MICB molecule.     

HLA-A*02:251新等位基因克隆测序鉴定及序列分析

目的 鉴定中国人群人类白细胞抗原(human leukocyte antigen,HLA)A*02:251新等位基因,分析新等位基因遗传特征.方法 采用聚合酶链反应-测序分型法(polymerase chain reaction-sequence based typing,PCR-SBT)对组织配型健康供、患者进行HLA基因分型,发现先证者核苷酸杂合序列与已知序列不匹配,不能指定先证者HLA等位基因型,对先证者DNA扩增HLA-A位点第2～4外显子,PCR产物经克隆到PMD18-T质粒载体中以获得单链核苷酸序列,对克隆所得产物进行HLA-A基因的第2～4外显子双向测序分析.结果 发现先证者的一个HLA-A*02:06:01基因被确认,而另一个HLA-A基因为新等位基因,其序列被GenBank接受(编号为HM245348).新等位基因序列通过IMGT/HLA 数据库BLAST,与最相近的A*02:01:01:01相比,在第3外显子上有1个核苷酸的不同,即第383位 G>C,密码子 128 GAG→GAC,氨基酸由谷氨酸(Glu)→天门冬氨酸(Asp).供、患者HLA-A、B、C、DQB1位点等位基因不匹配.结论 该等位基因为新的HLA-A*02:251等位基因.中国人群HLA-A 位点第3外显子核苷酸序列存在多态性.

Abstract：

Objective To identify a novel human leukocyte antigen (HLA) allele A*02:251 and analyze the sequences in Chinese population. Methods Routine HLA-A, -B, -DRB1 high resolution genotyping for healthy Chinese donors and patients was performed with polymerase chain reaction-sequence based typing. An unknown HLA-A allele was initially detected by HLA typing in the healthy donor. Genomic DNA of the HLA-A locus in the proband was amplified, the amplified product was cloned by PMD18-T to split the two alleles, and selected clones were sequenced. Results The sequencing results showed that a normal A*02:06:01 and a novel A*02:251 variant allele were identified. The sequence of the novel allele has been submitted to GenBank (HM245348). Nucleotide sequence alignments with HLA-A allele from the IMGT/HLA Sequence Database showed that the novel A*02 variant allele differed from the closest allele A*02:01:01:01 by nt 383 G>C (codon 128 GAG>GAC) in exon 3, which resulted in one amino acid substitution of Glu>Asp. The HLA-A, B, C and DQB1 alleles of the healthy donor did not match with that of the patient. Conclusion This novel allele is officially designated as HLA-A*02:251 by World Health Organization(WHO) Nomenclature Committee (Submission ID HWS10010755). The sequence of HLA-A locus in exon 3 is confirmed to be polymorphic in Chinese population.     

HLA‐B allele dropout in PCR sequence‐specific oligonucleotide probe typing due to intronic polymorphism in the novel B*58:01:01:02 allele

Currently, Luminex technology based on the PCR sequence‐specific oligonucleotide (SSO) probe method has been widely used for HLA genotyping in the immunogenetics laboratories. Here, we reported a case with HLA‐B allele dropout by Luminex technology. The initial HLA‐B result of the Luminex method with a commercial agent kit was inconclusive, and then, the result of PCR‐SBT technology indicated the dropout as a HLA‐B*58 allele. Subsequently, the full‐length sequence of HLA‐B allele was determined by TOPO‐TA cloning, and a novel allele B*58:01:01:02 was identified in the individual. Compared with HLA‐B*58:01:01:01, the novel allele showed some nucleotides difference at 509 C>T, 521 T>G and CCC insertion in position 503 of intron 2. According to the full‐length sequence, the new mutations of intron 2 were contributed to HLA‐B locus allele dropout in the sample. Our results indicated multiplatform should be used to improve the HLA typing accuracy when a conclusive HLA genotype cannot be determined.     

Identification of a novel HLA-C*02 allele, HLA-C*02:02:09

We report HLA-C*02:02:09 as a novel allele with a transition C->T at position 354.     

A novel HLA‐B*14 allele – B*14:53 – genetics and serology

HLA‐B*14:53 was found in a UK European normal blood donor prior to registration on the Welsh Bone Marrow Donor Registry. It differs from B*14:13 by one base (103G>T) in exon 2 resulting in a substitution of alanine (A) in B*14:13 to serine (S) in B*14:53. Unique among current HLA‐B*14 alleles, B*14:53 and B*14:13 share a motif of 59 bases between positions 361 and 419 in exon 3. This motif is present in numerous HLA‐B alleles the commonest overall being B*08:01, suggesting that both B*14:53 and B*14:13 arose from intralocus gene conversion events with B*08:01. Thus, B*14:53 probably arose from B*14:01:01 (which has TCC at codon 11 (S), while B*14:13 arose from B*14:02:01:01 which has GCC at codon 11 (A). Additionally, the two likely B*14:53‐bearing and B*14:13‐bearing haplotypes are typical of B*14:01:01‐bearing and B*14:02:01:01‐bearing haplotypes, respectively. Serological testing, using 49 antisera with HLA‐B64, or B64, B65 reactivity, showed that the B*14:53 specificity did not react as a B64 (B*14:01) specificity and may appear as a short/weak HLA‐B14. This implies that residues additional to S at position 11 are involved in HLA‐B64 serological identity; for example, the motif 11S 97W 116F is possessed by B*14:01 and many other B*14 products (and B*39:79 plus some HLA‐C products) but not B65 (B*14:02) or the B*14:53 specificity. B*14:53 was found in a random HLA sequence‐based typed population of 32 530 normal subjects indicating a low precision allele frequency of 0.000015 in subjects resident in Wales.