汉族人群中22个HLA-Cw等位基因全长序列单核苷酸多态性分析

为探讨HLA-Cw (Human leukocyte antigen-Cw)基因全长序列分子遗传多态性, 文章对28个HLA-Cw基因型已知的汉族个体样本, 采用长距离PCR技术和高保真性的Pfu酶, 扩增HLA-Cw基因全长序列4.5 kb, 进行分子克隆和单倍体测序。采用群体遗传学研究方法分析了HLA-Cw等位基因全长序列中各亚区的单核苷酸多态性。结果表明: 在28个样本中共检测出22种等位基因, 序列均已提交GenBank和国际IMGT/HLA数据库并获得了认可; 其中Cw*0706、Cw*030301、Cw*140201的全长序列为首次报道, 尤其是Cw*0706内含子序列的获得, 能够重新设计对该等位基因测序分型的引物, 避免测序分型中可能对这一等位基因的漏检。将28个样本的56条单倍体序列用Clustal软件进行序列排比, 输入到DnaSP4.0进行多态性分析, 共发现244个SNPs, 10处插入/缺失多态性。对HLA-Cw等位基因各亚区多态性的分析, 发现第4内含子及以前并没有受到关注的第5外显子受到平衡选择的作用, 在进化中受到了选择压力, 预示着它们在免疫系统的进化过程中可能扮演着重要的角色。     

内蒙古地区蒙古族HLA-A、B、DRB1基因座多态性分析

应用序列特异性寡核苷酸探针反向斑点杂交技术对内蒙古地区蒙古族106名无关健康个体的HLA-A、B和DRB1 基因座进行基因分型, 以研究内蒙古地区蒙古族人群HLA-A、B、DRB1基因座的等位基因及其组成的单倍型频率分布特征。 采用最大数学预期值算法计算HLA基因座的等位基因频率和单倍型频率。106 名内蒙古地区蒙古族个体的HLA-A、B、DRB1基因座分别检出13、29、13个等位基因。高频单倍型分别为 HLA-A*02-B*46 (0.0510); HLA-A*02-B*13(0.0495); HLA-A*02-B*51(0.0442); HLA-B*13-DRB1*07 (0.0555); HLA- B*46-DRB1*09(0.0378); HLA-B*35-DRB1*13(0.03300); HLA-A*02-B*13-DRB1*07(0.033019); HLA-A*02-B*46- DRB1*09(0.031985)。研究表明: 内蒙古地区蒙古族人群HLA基因座的等位基因和单倍型具有较高的遗传多态性。HLA- A*24-B*14, HLA-A*32-B*63在该民族具有极强的连锁不平衡。     

稀有的黑檀体基因（ebony）5′UTR自发突变导致黑腹果蝇的黑条体突变

由于果蝇Drosophila群体中有很多自发突变其中包括多种体色突变, 因此它是一个研究自发突变的优秀的模式体系。本研究证实我们实验室发现的一个可以引起果蝇体色突变的自发突变（bsr）是一个黑檀体（e）的等位基因, 将其命名为e^bsr。序列分析显示e^bsr的5′端缺失了953个碱基, 其中包括外显子1后端的206个碱基及相连的内含子1的747个碱基。逆转录PCR结果显示5′端的缺失导致内含子1不能从mRNA中剪接掉, 由此导致该mRNA的翻译起始密码子AUG前端增加了一个3.2 kb的序列。该序列导致e^bsr的mRNA的5′UTR（5′-untranslated region）区较野生型基因增加近3 kb的长度。通过mRNA二级结构分析发现这个增加的3 kb的片段可以形成复杂的颈环结构（stem-loop）。免疫印迹结果显示该突变基因没有基因产物产生。本研究进一步证实了由于mRNA的5′UTR序列结构的改变可以影响到蛋白质的翻译。     

HLA-B*2736等位基因的序列分析

使用FLOW-SSO、PCR-SSP以及测序等分型技术, 发现一个与HLA-B*270401基因相关的未知基因。设计基因特异性引物单独扩增B*27基因的外显子2-5, 包括内含子2-4, 并进行双向测序, 分析与B*270401基因序列的差异。该基因的扩增产物为1 815 bp。与B*270401相比在外显子3和4共有10个碱基的改变, 从而使相应氨基酸发生错义或同义突变。碱基634 A→C (密码子130丝氨酸→精氨酸); 670 A→T (密码子142苏氨酸→丝氨酸); 683 G→T (密码子146色氨酸→亮氨酸); 698 A→T (密码子151谷氨酸→缬氨酸); 774 G→C (密码子176谷氨酸→天冬氨酸); 776 C→A (密码子177苏氨酸→赖氨酸); 781 C→G (密码子179谷氨酰胺→谷氨酸); 789 G→T (密码子181丙氨酸同义突变); 1 438 C→T (密码子206甘氨酸同义突变); 1 449 G→C (密码子210甘氨酸→丙氨酸)。在IMGT/HLA数据库中B*27组只有3个基因（B*270502 / 2706 / 2732）提交了内含子序列。该未知基因的内含子2序列与B*2706相同, 显示了与B*27组基因的同源性, 但其同源性在内含子3、4均未得到支持, 与B*27组基因相比, 内含子3的第106个碱基C→G, 碱基168缺失, 碱基179 G→A, 碱基536 G→A; 内含子4中碱基82 T→C。但其内含子3、4序列却与B*070201完全相同。该基因序列已提交GenBank, 编号为被DQ915176, 被WHO确认为HLA-B*2736等位基因。     

癫痫相关基因SCN1A启动子区多态性位点的生物信息学分析

SCN1A是多种神经系统疾病的致病基因,该基因的精确表达对于维持神经系统正常功能非常重要.为了认识SCN1A启动子区多态性位点(single nucleotide polymorphisms,SNPs)的保守性及其功能意义,应用生物信息学方法分析了目前已发表位于SCN1A启动子区的11个SNPs,分别命名S1～S11.分析结果表明,S3、S5和S7等位基因频率没有人种差异性,其余SNP等位基因频率均有人种差异性;接近核心启动子的SNPs要比远离核心启动子SNPs的保守程度高,提示靠近核心启动子的SNPs影响SCN1A基因表达的概率可能越大;大部分SNPs祖传等位基因在哺乳动物中是保守的(S3除外),暗示这些SNPs新生等位基因有可能在人类进化过程起到一定的作用:启动子分析软件预测发现,含S2、S4、S8及S9等4个SNPs不同等位基因的同一序列分别存在不同转录因子结合元件,而含S1和S11不同等位基因的同一序列都只能预测到含其中一个等位基因的序列存在转录因子结合元件,这些差异可能是SNPs影响SCN1A表达的重要原因之一.这些分析将为进一步研究SCN1A启动子区SNPs与神经系统疾病的相互关系打下基础.     

三角帆蚌GPX基因结构特征及抗性相关SNP的筛选

根据三角帆蚌(Hyriopsis cumingii)谷胱甘肽过氧化物酶(Glutathione peroxidase,GPX)基因cDNA序列,通过PCR和基因组步移法,从三角帆蚌基因组DNA中扩增出GPX基因全长及其5′调控区。序列分析表明,该基因序列全长6 708 bp,含有2个外显子,1个内含子。5′调控区为992 bp,含有启动子的核心序列TATA盒以及其他一些转录调控元件,如AP1、C/EBP、CdxA。2个外显子长度分别为273 bp和991 bp,内含子长度为4 491 bp。通过直接测序法在三角帆蚌抗性群体和易感群体中筛选GPX基因的SNPs,并研究这些多态性位点与抗逆性状的相关性。共获得了16个SNP位点,其中启动子区的A-99G位点、A-86C位点、A-49C位点,内含子区的A2841T位点、C2847T位点、G3146C位点、A3150G位点以及G4645T位点共8个SNP位点的基因型频率和等位基因频率在抗性和易感群体中均存在显著性差异(P<0.05)。连锁不平衡分析结果显示GPX基因A-86C位点、A-49C位点、C2847T位点、A3150G位点与G4645T位点之间,以及A2841T位点与G3146C位点之间均存在强连锁不平衡。同时单倍型分析发现,在抗性群体中单倍型分别为ACTGT和TG的个体出现的频率显著高于易感群体中出现的频率。这些结果表明,GPX基因的部分SNPs可作为三角帆蚌抗病辅助育种的候选分子标记。     

猪CAPN1基因部分外显子及3′UTR区的SNPs检测

以野猪、民猪和大白猪为研究对象, 根据网上公布的序列设计了7对引物, 采用测序、PCR-SSCP和PCR-RFLP方法对CAPN1基因的部分外显子和3′UTR区进行了单核苷酸多态性检测和基因型分析, 探讨CAPN1基因多态性与瘦肉率和嫩度的关系。研究发现11个SNPs, 其中5个位于外显子, 4个位于内含子, 2个位于3′UTR区, 外显子中的突变有一处是错义突变, 导致了蛋白质多肽链第260位氨基酸发生了M/V的替代。群体遗传学分析表明, 在所检测的各多态位点上, 野猪、民猪、大白猪3个品种间不同基因型的分布都存在着极显著的差异(P<0.01), 而野猪和民猪之间各基因型的分布差异不显著(P>0.05), 民猪和大白猪之间各基因型的分布存在着极显著的差异(P<0.01)。结合品种特性分析表明, P4、P6引物和3′ UTR区HinfⅠ位点所检测的不同基因型和瘦肉率具有一定的相关性。     

中华绒螯蟹蜕皮抑制激素1(Ers-MIH1)基因组DNA的分子克隆和序列分析

运用反向PCR (IPCR)技术首次克隆得到全长为 3 50 6bp的中华绒螯蟹 (Eriocheirjaponicasinensis)蜕皮抑制激素 1(MIH 1)基因组DNA序列 (GenBank检索号 :AY3 10 3 13 )。该序列包括 3个外显子、 2个内含子、 412bp的 5′端上游调控区和 917bp的 3′端UTR。编码区的第 1个内含子将信号肽分开 ,第 2个内含子将成熟肽分开。MIH 1基因的外显子和内含子接头区符合受体拼接点和供体拼接点的GT AG法则。MIH 1基因412bp的 5′端侧翼区含有和其它真核基因相似的启动子元件 ,即包括与其它节肢动物高度相似的起始子、TATA盒以及cAMP效应元件结合蛋白的结合位点序列。中华绒螯蟹MIH 1基因的组织方式与斑纹和食用黄道蟹的MIH基因相同。推导的多肽由 75个氨基酸的成熟肽和 3 5个氨基酸的信号肽组成 ,成熟肽的氨基酸序列和食用黄道蟹、三叶真蟹及美洲黄道蟹的一致性在 64% -65%之间     

雌激素受体2（ESR2）mRNA 5′非翻译区内部核糖体进入位点活性的鉴定与分析

真核生物除了传统的帽依赖型翻译机制外,还存在内部核糖体进入位点（internal ribosome entry site, IRES）介导的翻译机制。雌激素受体2（estrogen receptor 2, ESR2）是雌激素受体家族成员之一,其编码的蛋白质在许多肿瘤中发挥重要的作用。ESR2蛋白的异常表达会导致众多肿瘤的发生,但其蛋白质翻译水平的调控机制至今仍不清楚。研究发现,在药物刺激的条件下,乳腺癌细胞MCF7/WT中ESR2蛋白的表达提高,但是其转录水平基本未见发生改变。猜测ESR2 mRNA 5′非翻译区（5′ untranslated region, 5′ UTR）具有IRES活性。为了验证ESR2 mRNA 5′ UTR是否具有IRES元件,将ESR2 mRNA 5′ UTR插入到双顺反子报告基因载体（pRF）中,构建pRL-ESR2-FL重组质粒载体,将其瞬时转染到HEK293细胞。结果发现,ESR2 mRNA 5′ UTR有假定的IRES活性。并且通过3个排除实验验证了ESR2 IRES活性与其5′ UTR中的内部潜在启动子（P<0.0001）、内部剪切位点以及核糖体通读无关。进一步对其序列进行截短研究发现,ESR2 IRES活性发挥的关键区域是3′端的439～468 nt,且ESR2 IRES最大活性的发挥依赖于5′ UTR序列的完整性。并且发现,ESR2 IRES活性的发挥不但需要特定的一级核酸序列,还要有稳定的二级茎环结构。此研究有望为ESR2蛋白调控的相关疾病提供新的药物治疗靶点。     

尼罗罗非鱼β2m基因SNP位点和单倍型与无乳链球菌抗性的关联分析

β₂-微球蛋白(β₂-microglobulin, β₂m)作为MHCⅠ类分子的亚基, 在鱼类的免疫系统中发挥重要作用。实验采用直接测序法从P₀代尼罗罗非鱼(Oreochromis niloticus)的β₂m基因组序列中筛选到30个SNPs, 其中1个SNP位于5?UTR, 16个SNPs位于外显子区域(15个非同义突变位点, 1个同义突变位点), 9个SNPs位于内含子区域, 4个SNPs位于3?UTR。利用snapshot分型法对F₁代的102尾易感群体和102尾抗病群体进行基因分型, 并通过Popgen32和PIC-CALC软件统计分析尼罗罗非鱼β₂m基因序列的SNPs的H_e、H_o、N_e和PIC等遗传参数, 表明易感群体中7个SNPs属于中度多态水平(0.251代2个群体中的基因型频率和等位基因频率, 分析其与链球菌抗性或易感性状之间的相关性。结果表明: 24个SNPs的基因型和等位基因频率与无乳链球菌(Streptococcus agalactiae)抗性/易感性状显著相关(P<0.05)。通过连锁不平衡分析发现30个SNPs构成4个单倍块和14种单倍型。其中, 4个单倍型与无乳链球菌抗性性状显著相关(P<0.05), 4个单倍型与易感性状显著相关(P<0.05)。标签SNP分析发现, 单倍块2中的4个SNPs和单倍块3中的13个SNPs彼此之间高度连锁(r2>0.9), 这意味着我们在β₂m基因中发现2个htSNPs。研究筛选到的与链球菌抗性/易感性状相关的SNP位点及单倍型具有辅助尼罗罗非鱼抗链球菌病品种选育的潜力。     

High Resolution Human Leukocyte Antigen Class I Allele Frequencies and HIV-1 Infection Associations in Chinese Han and Uyghur Cohorts

Background

Host immunogenetic factors such as HLA class I polymorphism are important to HIV-1 infection risk and AIDS progression. Previous studies using high-resolution HLA class I profile data of Chinese populations appeared insufficient to provide information for HIV-1 vaccine development and clinical trial design. Here we reported HLA class I association with HIV-1 susceptibility in a Chinese Han and a Chinese Uyghur cohort.

Methodology/Principal Findings

Our cohort included 327 Han and 161 Uyghur ethnic individuals. Each cohort included HIV-1 seropositive and HIV-1 seronegative subjects. Four-digit HLA class I typing was performed by sequencing-based typing and high-resolution PCR-sequence specific primer. We compared the HLA class I allele and inferred haplotype frequencies between HIV-1 seropositive and seronegative groups. A neighbor-joining tree between our cohorts and other populations was constructed based on allele frequencies of HLA-A and HLA-B loci. We identified 58 HLA-A, 75 HLA-B, and 32 HLA-Cw distinct alleles from our cohort and no novel alleles. The frequency of HLA-B*5201 and A*0301 was significantly higher in the Han HIV-1 negative group. The frequency of HLA-B*5101 was significantly higher in the Uyghur HIV-1 negative group. We observed statistically significant increases in expectation-maximization (EM) algorithm predicted haplotype frequencies of HLA-A*0201-B*5101 in the Uyghur HIV-1 negative group, and of Cw*0304-B*4001 in the Han HIV-1 negative group. The B62s supertype frequency was found to be significantly higher in the Han HIV-1 negative group than in the Han HIV-1 positive group.

Conclusions

At the four-digit level, several HLA class I alleles and haplotypes were associated with lower HIV-1 susceptibility. Homogeneity of HLA class I and Bw4/Bw6 heterozygosity were not associated with HIV-1 susceptibility in our cohort. These observations contribute to the Chinese HLA database and could prove useful in the development of HIV-1 vaccine candidates.     

Preferential HLA usage in the influenza virus-specific CTL response

To study whether individual HLA class I alleles are used preferentially or equally in human virus-specific CTL responses, the contribution of individual HLA-A and -B alleles to the human influenza virus-specific CTL response was investigated. To this end, PBMC were obtained from three groups of HLA-A and -B identical blood donors and stimulated with influenza virus. In the virus-specific CD8(+) T cell population, the proportion of IFN-gamma- and TNF-alpha-producing cells, restricted by individual HLA-A and -B alleles, was determined using virus-infected C1R cells expressing a single HLA-A or -B allele for restimulation of these cells. In HLA-B*2705- and HLA-B*3501-positive individuals, these alleles were preferentially used in the influenza A virus-specific CTL response, while the contribution of HLA-B*0801 and HLA-A*0101 was minor in these donors. The magnitude of the HLA-B*0801-restricted response was even lower in the presence of HLA-B*2705. C1R cells expressing HLA-B*2705, HLA-A*0101, or HLA-A*0201 were preferentially lysed by virus-specific CD8(+) T cells. In contrast, the CTL response to influenza B virus was mainly directed toward HLA-B*0801-restricted epitopes. Thus, the preferential use of HLA alleles depended on the virus studied.     

HLA and non-HLA genes in Beh?et’s disease: a multicentric study in the Spanish population

Introduction

According to genome wide association (GWA) studies as well as candidate gene approaches, Behçet’s disease (BD) is associated with human leukocyte antigen (HLA)-A and HLA-B gene regions. The HLA-B51 has been consistently associated with the disease, but the role of other HLA class I molecules remains controversial. Recently, variants in non-HLA genes have also been associated with BD. The aims of this study were to further investigate the influence of the HLA region in BD and to explore the relationship with non-HLA genes recently described to be associated in other populations.

Methods

This study included 304 BD patients and 313 ethnically matched controls. HLA-A and HLA-B low resolution typing was carried out by PCR-SSOP Luminex. Eleven tag single nucleotide polymorphisms (SNPs) located outside of the HLA-region, previously described associated with the disease in GWA studies and having a minor allele frequency in Caucasians greater than 0.15 were genotyped using TaqMan assays. Phenotypic and genotypic frequencies were estimated by direct counting and distributions were compared using the χ² test.

Results

In addition to HLA-B*51, HLA-B*57 was found as a risk factor in BD, whereas, B*35 was found to be protective. Other HLA-A and B specificities were suggestive of association with the disease as risk (A*02 and A*24) or protective factors (A*03 and B*58). Regarding the non-HLA genes, the three SNPs located in IL23R and one of the SNPs in IL10 were found to be significantly associated with susceptibility to BD in our population.

Conclusion

Different HLA specificities are associated with Behçet’s disease in addition to B*51. Other non-HLA genes, such as IL23R and IL-10, play a role in the susceptibility to the disease.     

MICA genetic polymorphism and linkage disequilibrium with HLA-B in 29 African-American families

The human major histocompatibility complex (MHC) class I chain-related gene A ( MICA) is located 46 kb upstream of HLA-B and encodes a stress-inducible protein which displays a restricted pattern of tissue expression. MICA molecules interact with NKG2D, augmenting the activation of natural killer cells, CD8(+) alpha beta T cells, and gamma delta T cells. MICA allelic variation is thought to be associated with disease susceptibility and immune response to transplants. We investigated MICA allelic variations and linkage disequilibrium with HLA-A, B, and DRB1 loci on 110 parental haplotypes from 29 African-American families. PCR/sequence-specific oligonucleotide probing (SSOP) was used to define MICA polymorphisms in exons 2, 3, and 4. Ambiguous allelic combinations were resolved by sequencing exons 2, 3, and 4. Exon 5 polymorphisms were analyzed by size sequencing. For HLA-A, B and DRB1 typing, low-resolution PCR/SSOP and allelic PCR/sequence-specific priming techniques were used. Twelve MICA alleles were observed, the most frequent of which were MICA*008, MICA*004, and MICA*002, with gene frequencies of 28.2, 26.4, and 25.5%, respectively. Thirty-eight HLA-B- MICA haplotypic combinations were uncovered, 22 of which have not been reported in the HLA homozygous typing cell lines from the 10th International Histocompatibility Workshop. Significant positive linkage disequilibria were found in 8 HLA-B- MICA haplotypes. Furthermore, haplotypes bearing HLA-B*1503, *1801, *4901, *5201, *5301, and *5703 were found to segregate with at least two different MICA alleles. Our results provide new data about MICA genetic polymorphisms in African-Americans, which will form the basis for future studies of MICA alleles in allogeneic stem cell transplantation outcome.     

Survival and HLA-B*57 in HIV/HCV Co-Infected Patients on Highly Active Antiretroviral Therapy (HAART)

Background and aims

HLA class I alleles, in particular HLA-B*57, constitute the most consistent host factor determining outcomes in untreated HCV- and HIV-infection. In this prospective cohort study, we analysed the impact of HLA class I alleles on all-cause mortality in patients with HIV-, HCV- and HIV/HCV- co-infection receiving HAART.

Methods

In 2003 HLA-A and B alleles were determined and patients were prospectively followed in 3-month intervals until 2013 or death. HLA-A and B alleles were determined by strand-specific oligonucleotide hybridisation and PCR in 468 Caucasian patients with HCV- (n=120), HIV- (n=186) and HIV/HCV-infection (n=162). All patients with HIV-infection were on HAART. In each patient group, HLA class I-associated survival was analysed by Kaplan-Meier method and Cox regression analysis.

Results

At recruitment the proportion of patients carrying a HLA-B*57 allele differed between HIV- (12.9%) and HCV-infection (4.2%). Kaplan Meier analysis revealed significantly increased mortality in HLA-B*57-positive patients with HIV-infection (p=0.032) and HIV/HCV-co-infection (p=0.004), which was apparently linked to non-viral infections. Cox logistic regression analysis confirmed HLA-B*57 (p=0.001), serum gamma-glutamyltranspeptidase (p=0.003), serum bilirubin (p=0.022) and CD4 counts (p=0.041) as independent predictors of death in HIV-infected patients.

Conclusion

Differences in the prevalence of HLA-B*57 at study entry between HIV- and HCV- infected patients may reflect immune selection in the absence of antiviral therapy. When patients were treated with HAART, however, HLA-B*57 was associated with increased mortality and risk to die from bacterial infections and sepsis, suggesting an ambiguous role of HLA-B*57 for survival in HIV/HCV infection depending on the circumstances.     

Diversity and diversification of HLA-A,B,C alleles

The nucleotide sequences encoding 14 HLA-A,B,C and 5 ChLA-A,B,C molecules have been determined. Combining these sequences with published data has enabled the polymorphism in 40 HLA-A,B,C and 9 ChLA-A,B,C alleles to be analyzed. Diversity is generated through assortment of point mutations by recombinational mechanisms including gene and allelic conversions. The distribution and frequency of silent and replacement substitutions indicate that there has been positive selection for allelic diversity in the 5' part of the gene (exons 1 to 3) and for allelic homogenization and locus specificity in the 3' part of the gene (exons 4 to 8). These differences may correlate with the lengths of converted sequences in the two parts of the gene and frequency of the CpG dinucleotide. Locus-specific divergence of HLA-A,B, and C demonstrates that recombinational events involving alleles of a locus have been more important than conversion between loci. This contrasts with the predominance of gene conversion events in the evolution of mutants of the H-2Kb gene. However, a striking example of gene conversion involving HLA-B and C alleles of an oriental haplotype has been found. Comparison of human and chimpanzee alleles reveals extensive sharing of polymorphisms, confirming that diversification is a slow process, and that much of contemporary polymorphism originated in ancestral primate species before the emergence of Homo sapiens. There is less polymorphism at the HLA-A locus compared to HLA-B, with greater similarity also being seen between HLA-A and ChLA-A alleles than between HLA-B and ChLA-B alleles. Although greater diversity is seen in the 5' "variable" exons of HLA-B compared to HLA-A, there is increased heterogeneity in the 3' "conserved" exons of HLA-A compared to HLA-B.     

The nature of introns 4-7 largely reflects the lineage specificity of HLA-A alleles

For most HLA-A alleles the phylogeny of the 3' non-coding regions has not yet been studied systematically. In this study, we have determined the sequences of introns 4-7 in 50 HLA-A variants, and have computed nucleotide substitution rates and phylogenetic relationships. The A2/A28, A9, and A10 groups were characterized by clear lineage specificity. For the A19 group, lineage specificity was weaker. A*3001 clustered together with the alleles of the A1/A3/A11/A36 serological family, but not with the A19 group alleles. Reduced lineage specificity was also observed for the alleles of the A1/A3/A11/A36groups. The 3' intron sequences of A*8001 were clearly distinct from all other alleles studied. In several cases two allelic groups shared identical intron sequences, whereby the patterns varied with the introns. A similar situation has been previously described for the 5' introns. Since recombination is the major mechanism of HLA diversification, the intronic lineage specificity corresponds to the comparatively lower recombination rate of the HLA-A 3' exons. The low level of recombination within the 3' region of HLA-A is supported by the low CpG content with a maximum of 3.0% in this region compared with up to 10.7% in the 5' region. Apart from phylogenetic studies of HLA diversity and diversification, the sequence data obtained in our study may prove valuable for the development of a haplotype-specific sequencing strategy for the HLA-A3' exons and for the explanation of recombination events in newly described HLA class I alleles.