The beta globin gene cluster of the prosimian primate Galago crassicaudatus: nucleotide sequence determination of the 41-kb cluster and comparative sequence analyses.

The nucleotide sequence of the beta globin gene cluster of the prosimian Galago crassicaudatus has been determined. A total sequence spanning 41,101 bp contains and links together previously published sequences of the five galago beta-like globin genes (5'-epsilon-gamma-psi eta-delta-beta-3'). A computer-aided search for middle interspersed repetitive sequences identified 10 LINE (L1) elements, including a 5' truncated repeat that is orthologous to the full-length L1 element found in the human epsilon-gamma intergenic region. SINE elements that were identified included one Alu type I repeat, four Alu type II repeats, and two methionine tRNA-derived Monomer (type III) elements. Alu type II and Monomer sequences are unique to the galago genome. Structural analyses of the cluster sequence reveals that it is relatively A+T rich (about 62%) and regions with high G+C content are associated primarily with globin coding regions. Comparative analyses with the beta globin cluster sequences of human, rabbit, and mouse reveal extensive sequence homologies in their genic regions, but only human, galago, and rabbit sequences share extensive intergenic sequence homologies. Divergence analyses of aligned intergenic and flanking sequences from orthologous human, galago, and rabbit sequences show a gradation in the rate of nucleotide sequence evolution along the cluster where sequences 5' of the epsilon globin gene region show the least sequence divergence and sequences just 5' of the beta globin gene region show the greatest sequence divergence.     

Structure and sequence of the mitochondrial 20S rRNA and tRNA tyr gene of Paramecium primaurelia

The sequence and structure of the large (20s) mitochondrial (mt) rRNA gene and flanking regions from Paramecium primaurelia have been determined. The gene contains two regions of strong homology with other large mt rRNAs: one 44-base region near the 5' end and a 321-base region near the 3' end. Another region of strong homology to both ends of E. coli 23s RNA exists at loci consistent with these regions. The Paramecium gene appears to be 2204 bases in length and contains slightly more homology to E. coli rRNA than its mammalian or fungal counterparts. The gene, located about 1200 bp from the replicative terminal end of the linear mt DNA, is transcribed in the same polarity as replication. Previous R-looping studies detected no large introns within the gene. Here we describe sequences resembling degenerate rRNAs, one of which could represent a small intron. A tRNA tyr gene was found on the same DNA strand, 127 bp downstream from the large rRNA presumptive 3' end. The tRNA is flanked on both sides by short DNA regions of approximately 90% A + T content.     

The structure of the human apolipoprotein C-II gene. Electron microscopic analysis of RNA:DNA hybrids, complete nucleotide sequence, and identification of 5' homologous sequences among apolipoprotein genes

Cloned human apo-C-II cDNA was used as a hybridization probe to identify the human apo-C-II gene in a genomic library constructed in our laboratory. The isolated apo-C-II DNA was studied both by electron microscopy and by direct sequence analysis. Ultrastructural morphological analysis of RNA-DNA hybrids revealed that the apo-C-II gene had complex structures because of regions of inverted complementary sequences in and around the gene forming stem-and-loop structures which interfere with the formation of stable RNA:DNA hybrids. Extensive morphological analysis revealed a minimum of 3 intervening sequences (IVS), and their lengths were measured. Direct sequence analysis of the cloned gene confirmed the presence of 3 IVS. There are 4 Alu type sequences in IVS-I. We sequenced 4340 nucleotides which include 545 nucleotides in the 5' flanking region, the entire gene which spans 3320 nucleotides, and 475 nucleotides in the 3' flanking region which also encompasses an additional Alu sequence. The 5' end of the gene was identified by primer extension and sequencing of the primer extended cDNA. Apo-C-II mRNA structure was deduced from the cDNA sequence, the primer extension experiments, and the genomic sequence. It is 494 nucleotides in length. Its sequence differs from previously published sequences in that there are 7 additional nucleotides before the polyadenylate tail. In the 5' flanking region, nucleotides -234 to -213 encompass a GC-rich region which exhibits high homology (greater than 70%) to the 5' flanking regions of the genes of all the apolipoproteins published to date, namely, apo-A-II (-497 to -471), apo-A-I (approximately -196 to -179), apo-E (-409 to -391), and apo-C-III (approximately -116 to -103). This highly conserved region might represent some evolutionarily conserved sequences from these related genes and/or might represent a region with regulatory function.     

Nucleotide sequence of the 3' nuclease-sensitive region of the human phosphoglycerate kinase 1 (PGK1) gene.

Many genes are known to have nuclease-sensitive sites and/or control sequences in their 3' flanking regions, but for very few genes has this region been sequenced. Previously, we mapped specific, gene activity-dependent DNAase I- and MspI-sensitive sites at the 3' end of the human X-linked housekeeping gene phosphoglycerate kinase (PGK1). Sequence information presented here shows that the 3' nuclease-sensitive site maps precisely to an Alu sequence and near a "BKM" repeat. This is the first report of an Alu sequence that has alternative chromatin configurations depending on gene activity.     

Isolation and complete nucleotide sequence of the gene for bovine parathyroid hormone

The structure of the bovine parathyroid hormone (PTH) gene has been analyzed by Southern blot hybridization of genomic DNA and by nucleotide sequence analysis of a cloned PTH gene. In the Southern analysis, several restriction enzymes produced single fragments that hybridized to PTH cDNA suggesting that there is a single bovine PTH gene. The restriction map of the cloned gene is the same as that determined by Southern blot analysis of bovine DNA. The sequence of 3154 bp of the cloned gene has been determined including 510 bp and 139 bp in the 5' and 3' flanking regions, respectively. The gene contains two introns which separate three exons that code primarily for: (i) the 5' untranslated region, (ii) the pre-sequence of preProPTH, and (iii) PTH and the 3' untranslated region. The gene contains 68% A + T and unusually long stretches of 100- to 150-bp sequences containing alternating A and T nucleotides in the 5' flanking region and intron A. The 5' flanking region contains two TATA sequences, both of which appear to be functional as determined by S1 nuclease mapping. Compared to the rat and human genes, the locations of the introns are identical but the sizes differ. Comparable human and bovine sequences in the flanking regions and introns are about 80% homologous.     

Molecular structure and flanking nucleotide sequences of the natural chicken ovomucoid gene

Five independent clones containing the natural chicken ovomucoid gene have been isolated from a chicken gene library. One of these clones, CL21, contains the complete ovomucoid gene and includes more than 3 kb of DNA sequences flanking both termini of the gene. Restriction endonuclease mapping, electron microscopy and direct DNA sequencing analyses of this clone have revealed that the ovomucoid gene is 5.6 kb long and codes for a messenger RNA of 821 nucleotides. The structural gene sequence coding Ifor the mature messenger RNA is split into at least eight segments by a minimum of seven intervening sequences of various sizes. The shortest structural gene segment is only 20 nucleotides long. All seven intervening sequences are located within the peptide coding region of the gene, and the sequences at the 5' and 3' untranslated regions of the mRNA are not interrupted by intervening sequences. The DNA sequences of the regions flanking the 5' and 3' termini of the gene have been determined. Thirty nucleotides before the start of the messenger RNA coding sequence is the heptanucleotide TATATAT, which is also present in a similar location relative to the chicken ovalbumin gene and other unique sequence eucaryotic genes. This sequence resembles that of the Pribnow box in procaryotic genes where a promoter function has been implicated. Seven nucleotides past the 3' end of the gene is the tetranucleotide TTGT, a sequence found to be present at identical locations as either TTTT or TTGT in other eucaryotic genes that have been sequenced. These conserved DNA sequences flanking eucaryotic genes may serve some regulator function in the expression of these genes.     

Gene Conversions Can Generate Sequence Variants in the Late Chorion Multigene Families of Bombyx Mori

The 140-kbp late chorion locus of Bombyx mori strain 703 contains 15 divergently oriented gene pairs encoding the high cysteine (Hc) eggshell proteins. Sequence homology is approximately 91% for the 2-kb region of each gene pair, including the 5' flanking region, intron and exons. The homology rapidly disappears within a few hundred basepairs of the 3' end of most genes. Here we present the results of the nucleotide sequence and genomic blot comparison of Hc genes from different races of B. mori. Comparison of the nucleotide sequences of the same gene pair in two different races reveals that most of the nucleotide differences occur in clusters or patches and correspond to sequences present in other Hc genes in the locus. The number of nucleotide differences that have accumulated in the highly conserved regions of the gene pair (2.3/100 bp), most of which are attributable to patchwork exchanges, is significantly higher than the number of differences in the poorly conserved 3' flanking regions (0.6/100 bp), due primarily to new mutations. These data are consistent with a gene conversion process, which in the short-term generates new combinations of sequence variants, but in the long-term results in concerted evolution. Genomic blot analyses of different geographical races of B. mori reveal that there is variation in the number of Hc gene pairs (14-19 gene pairs), indicating that unequal crossovers also occur in the locus.