A silent deletion in the beta-globin gene cluster.

A survey of the gamma-globin gene region of over 1000 normal individuals revealed a novel 2.5 kb deletion which removes the 5' end of the A gamma-globin gene. Unusually, this deletion in the beta-globin gene cluster is not associated with increased fetal haemoglobin production. Sequence analysis of the deletion endpoints revealed no significant homology at the breakpoint and failed to support a role for a proposed recombination hotspot in IVS-2 in the generation of this illegitimate recombination event. The existence of small "silent" deletions in the beta-globin gene cluster emphasizes the importance of deletion size in altering expression of the fetal globin genes.     

Restriction site polymorphisms in the pig beta-globin gene cluster

A restriction fragment length polymorphism was detected in pig DNA digested with Hind III restriction endonuclease and probed with rabbit beta 1-globin gene. Eight different phenotypes were observed and for six of them family data demonstrated that they are determined by three alleles. As this polymorphism is not found with four other restriction endonucleases (Bam HI, Eco RI, Kpn I, and Pst I), single point mutations are proposed to explain the observed differences.     

The nucleotide sequence of the human beta-globin gene

We report the complete nucleotide sequence of the human beta-globin gene. The purpose of this study is to obtain information necessary to study the evolutionary relationships between members of the human beta-like globin gene family and to provide the basis for comparing normal beta-globin genes with those obtained from the DNA of individuals with genetic defects in hemoglobin expression.     

Nonuniform recombination within the human beta-globin gene cluster.

Population genetic analysis of 15 restriction site polymorphisms demonstrates nonuniform recombination within the human beta-globin gene cluster. These DNA polymorphisms show two clusters of high nonrandom associations, one 5' and another 3' to the beta-globin structural gene, with no significant linkage disequilibrium between the two clusters. The 5'- and 3'-association clusters are 34.6 kilobases (kb) and 19.4 kb long, respectively, and are separated by 9.1 kb of DNA immediately 5' to the beta-globin gene. For each of these three DNA regions, we have observed a relationship between nonrandom associations and physical distance between the polymorphisms. However, this relationship differed for each of these regions. On the assumption that the effective population size (Ne) is 5,000-50,000, we estimate the total recombination rate to be 0.0017%-0.0002% in the 5' cluster, 0.0931%-0.0093% in the 3' cluster, and 0.2912%-0.0219% in the 9.1-kb region between them. The beta cluster thus shows nonuniformity in recombination. Moreover, the recombination rate in the 9.1-kb DNA segment is 3-30 times greater than expected and is thus a hot spot for meiotic recombination.     

Detection of a novel DNA polymorphism in the beta-globin gene cluster

Analysis of DNA from the beta-globin gene cluster in an Albanian family identified a novel RsaI site approximately 550 base pairs 5' to the beta-globin gene. In this family, two chromosomes carrying otherwise identical beta-globin haplotypes were found to differ at the RsaI site. Population screening demonstrated the presence and absence of the site in DNA from individuals of northern European, Mediterranean, Middle Eastern, Southeast Asian, African, and Asian Indian descent, indicating that this site is a DNA polymorphism common in many ethnic groups. The polymorphism is also present in DNA from individuals carrying different beta-globin alleles. Additional nucleotide sequence changes identified in an RsaI (+) genomic clone in the region immediately 3' to the RsaI site suggest a mechanism for the randomization of the site with respect to haplotype.     

Nucleotide sequence of the beta-globin genes in gorilla and macaque: The origin of nucleotide polymorphisms in human

Summary Part of the beta-globin genes ofMacaca cynomolgus andGorilla gorilla has been cloned and sequenced. Ten putatively neutral nucleotide polymorphisms have been described at the beta-globin locus in humans. They are associated in seven combinations, which define seven different haplotypes of the beta-globin gene: four major frameworks—1, 2, 3, and 3^*—and three minor frameworks, which we term KI1, KA1, and OR1. The nucleotide sequences of these frameworks are compared with those of homologous sequences in chimpanzee, colobus, macaque, and gorilla. This comparison provides strong evidence that framework 2 was the earliest framework in the human lineage. From framework 2, a rooted parsimonious tree for the six other frameworks is constructed. This phylogenetic tree is discussed in terms of the evolution of nucleotide polymorphisms as well as in terms of genetic affinities between human populations.For each position at which there is base difference in comparing human, gorilla, and chimpanzee beta-globin genes, the phyletic lineage where the corresponding substitution occurred has been identified using the maximum parsimony procedure. The data provide evidence that polymorphisms may represent a significant component of differences between closely related species. If so, nucleotide polymorphisms may strongly bias estimates of small evolutionary distances.     

Site-specific transfer of an intact beta-globin gene cluster through a new targeting vector

The ideal gene-therapy vector for treating genetic disorders should deliver intact therapeutic genes and their essential regulatory elements into the specific "safe genomic site" and realize long-term, self-regulatory expression. For beta-thalassemia gene therapy, viral vectors have been broadly used, but the accompanying insertional mutation and immunogenicity remain problematic. Hence, we aimed to develop new non-viral vectors that are efficient and safe in treating diseases. As previous studies have demonstrated that physiological expression of beta-globin genes requires both a 5' locus control region and 3' specific elements, we constructed a new human chromosome-derived targeting vector to transfer the intact beta-globin gene cluster into K562 cells. The whole beta-globin gene cluster was precisely integrated into the target site and expressed in a self-regulatory pattern. The results proved that the human chromosome-derived vector was specifically targeted to the human genome and this could provide a novel platform for further gene therapy research.     

Spontaneous amplification of the actinorhodin gene cluster in Streptomyces coelicolor involving native insertion sequence IS466

We observed a spontaneous amplification of the Streptomyces coelicolor chromosome, including genes encoding biosynthetic enzymes of the antibiotic actinorhodin. A new junction of two tandem segments has, inserted within it, a third copy of a transposable element existing in two places elsewhere in the chromosome, suggesting its involvement in the amplification mechanism.     

The beta-globin gene cluster distribution revisited-patterns in Native American populations

New frequencies for the beta-globin gene cluster haplotypes are presented for the Aché (N = 82 individuals), Guarani (N = 76), and Kaingang (N = 54), three Native South American populations that live in an area between parallels 20 degrees S and 30 degrees S not covered by previous studies at this locus. The haplotype frequencies obtained for the three populations are within the interval observed for 28 other Native American populations. The Aché show much less haplotypes (five) than the other two populations (9-10), the haplotype prevalences being more similar to those of the Guarani than to the Kaingang. The Native American total heterozygosity was about half (0.41) that obtained for the African populations (0.71), but was not much different from those obtained for other continents. A geographical pattern was disclosed in South America by mapping the frequencies of the most common haplotype (haplotype 2), and by means of spatial correlation analysis. The analysis of molecular variance (AMOVA) and pairwise F(ST) data suggest three distinct sectors for the genetic landscape of Native South America: the Andes, the Center/Southeast region, and the Amazon.     

Nucleotide sequence analysis of the delta beta-globin gene region in humans

The continuous DNA sequence of a 16.5-kilobase pair region encompassing the linked delta beta-globin gene cluster in humans is presented with a detailed restriction endonuclease map. There are 38 differences (0.5%) in comparison with published sequence data, corrected for errors in sequencing, resulting in polymorphic rates of 0.2% in exons and 0.76% in 5'-gene flanking regions. Fifteen changes result in the generation or elimination of restriction sites which may be useful in linkage disequilibrium studies. Two pairs of inverted Alu repeats, a pyrimidine-rich region 5' to delta, and (TG)n, (Pu/Py)n, and (ATTTT)n tracts 5' to beta are described. Dinucleotide frequencies and deviation from expected values approximated those found in total human genomic DNA. Regions of less than 50% A + T content were found associated with Alu sequences, a 150-base pair region immediately 5' to the beta gene, exon regions from both genes, and an area 3' to the beta gene. These regions also contained significantly lower than expected CpG levels compared to other regions, suggesting a possible relationship between DNA organizational patterns and functionally important regions. In addition, strand asymmetries in base composition in this region differ from those associated with the fetal globin genes.     

Mitochondrial DNA sequence from an enigmatic gorilla population (Gorilla gorilla uellensis)

Although today gorillas are found in only two widely separate, discontinuous western and eastern African populations, rumors of the existence of an additional gorilla population in central Africa have inspired recent unsuccessful field expeditions in search of the "mystery ape" termed Gorilla gorilla uellensis. Such a gorilla population would have considerable conservation and scientific interest, and would presumably have descended from a population of gorillas that was thought to exist until the end of the 19th century on the Uele River in the current-day Democratic Republic of Congo. However, the sole evidence for the existence of these gorillas is three skulls and one mandible brought to the Royal Museum for Central Africa (Tervuren, Belgium) in 1898. We determined a mitochondrial DNA sequence from one of these specimens and compared it to sequences from other gorillas. Contrary to expectations, the sequence obtained did not exhibit the phylogenetic distinctiveness typical of a representative of a peripheral isolated population. Rather, the results suggest a scenario in which the museum specimens did not originally derive from the northern Congo, but were brought from the area of current distribution of western gorillas to that location; the subsequent discovery and collection of the specimens there gave rise to the false inference of a local gorilla population.     

Human 7SL RNA consists of a 140 nucleotide middle-repetitive sequence inserted in an alu sequence

We have cloned and sequenced a cDNA copy of in vitro-polyadenylated 7SL RNA of HeLa cells. The cloned fragment is 303 bp long and has a composite structure. A central block of 140 bp is homologous to a new set of human middle-repetitive sequences. This block appears to be inserted in an Alu consensus sequence, 100 bp from the 5' end and 40 bp from the 3' end of the Alu monomer. Two 6 bp direct repeats are found at the junction between the Alu flanking sequences and the central element. The analysis of several clones shows the existence of sequence microheterogeneity in the 5' portion of the molecule. The 7L DNA probably represents a subset of the Alu family of DNA, highly conserved in evolution.     

Insertion of a rabbit beta-globin gene sequence into an E. coli plasmid.

Double stranded DNA has been synthesized in vitro from rabbit globin messenger RNA and elongated with homopolymeric dG tails. An E. coli plasmid was cleaved by EcoRI. The cohesive ends were repaired and dC tails added, to permit reconstitution of the EcoRI sites upon annealing with the dG elongated globin DNA. Transformation of E. coli with the globin-plasmid DNA hybrid has yielded a clone which harbours a recombinant plasmid (pCR1-betaG1), as demonstrated by hybridization experiments with radioactive globin cDNA. The sequence carried by the recombinant plasmid corresponds to part of the gene sequence coding for the beta chain of rabbit globin. Circular DNA of the purified recombinant plasmid exhibits sensitivity to EcoRI.