Gene duplications and sequence polymorphism of bovine class II DQB genes

The genetic diversity of bovine class II DQB genes was investigated by polymerase chain reaction amplification and DNA sequencing. The first domain exon was amplified from genomic DNA samples representing 14 class II haplotypes, defined by restriction fragment length polymorphism (RFLP) analysis. The presence of a polymorphism in the copy number of DQB genes was confirmed since two DQB sequences were isolated from certain haplotypes. Four subtypes of bovine DQB genes were found. DQB1 is the major type and was found in almost all haplotypes. DQB2 is very similar to DQB1 but was found only in the duplicated haplotypes DQ9 to 12. DQB3 and DQB4 are two quite divergent genes only present in certain duplicated haplotypes. The bovine DQB complexity thus resembles that in the human DRB region. Bovine DQB genes were found to be highly polymorphic as ten DQB1 alleles and four DQB2 alleles were identified. The observed sequence polymorphism correlated well with previously defined DQB RFLPs. Bovine and human DQB alleles show striking similarities at the amino acid level. In contrast, the frequency of silent substitutions is much higher in comparisons of DQB alleles between species than within species ruling out the possibility that any of the contemporary DQB alleles have been maintained since the divergence of humans and cattle. The frequency of silent substitutions between DQB alleles was markedly lower in cattle than in humans, in agreement with a previous comparison of human and bovine DRB alleles.     

Genetic polymorphisms at the human liver/islet glucose transporter (GLUT2) gene locus in Caucasian and West Indian subjects with type 2 (non-insulin-dependent) diabetes mellitus.

The liver/islet glucose transporter (GLUT2) is mainly expressed in the hepatocytes of the liver and the beta-cells of the pancreatic islets and a defect in this transporter could lead to diabetic phenotypes, such as relative hypoinsulinaemia and reduced uptake and metabolism of glucose in the liver. DNA from unrelated individuals was digested with the restriction endonucleases Bgl-I and Taq-I followed by blotting and hybridisation with a 32P-labelled GLUT2 cDNA which revealed three restriction fragment length polymorphisms (RFLPs) (B1, T1 and T2) in both Caucasian and West Indian populations. Linkage analysis between these variant sites demonstrated that the alleles of these polymorphisms were in strong linkage disequilibrium. Disease association of genetic variants at the GLUT2 locus with type 2 diabetes was examined with these RFLPs in both Caucasian (n = 54) and West Indian (n = 46) populations with type 2 diabetes. There were no significant differences in the frequency of alleles, genotypes or haplotypes between diabetic patients and non-diabetic controls. However, there were significant differences in the allele frequencies of all these three polymorphisms between Caucasian and West Indian populations.     

Mucopolysaccharidosis IVA: polymorphic haplotypes and informative RFLPs in the Japanese population

Seven different restriction fragment length polymorphisms (RFLPs) at the N-acetylgalactosamine-6-sulfate sulfatase (GALNS) locus were analyzed using Southern blotting and polymerase chain reaction based techniques to search for the frequency of each RFLP produced by StyI, SphI, HaeIII, StuI, HapII, XhoI, and BamHI restriction endonucleases, respectively, in 36 mutant alleles, including two sibling cases and 100 normal alleles. Calculation of heterozygosity indexes showed that these RFLPs were polymorphic, ranging from 0.31 to 0.69 in mucopolysaccharidosis IVA (MPS IVA) patients compared with 0.21 to 0.65 in normal individuals. There was some significant difference in several RFLPs and in the combination with four kinds of RFLPs (SphI, StuI, HapII, XhoI polymorphisms). The normal alleles were composed of 13 different RFLPs haplotypes; the most common among the Japanese population carrying normal alleles was haplotype 8 (bDEF1) (31.3%), the others being dispersed. The same haplotype 8 was the most frequent in the mutant alleles (44.4%), with seven further haplotypes. These findings revealed the striking variety of polymorphic haplotypes in the MPS IVA gene. By using these five kinds of RFLPs, we examined the theoretical informativity of haplotype analysis in heterozygote detection in nine unrelated MPS IVA families and ten unrelated normal families. All the members of the MPS IVA families studied were diagnosed as a patient, carrier, or noncarrier. We propose that prenatal diagnosis or family analysis in cases in which mutations have not been characterized is now feasible.     

Multiple restriction fragment length polymorphisms in the porcine calcium release channel gene (CRC): assignment to the halothane (HAL) linkage group

Two cDNA probes for the porcine calcium release channel gene (CRC) were used in restriction fragment length polymorphism (RFLP) analysis in an attempt to develop genetic markers linked to the malignant hyperthermia (stress susceptibility) gene (HAL). Three TaqI RFLPs, denoted CRC1-CRC3, each composed of two alleles, were detected. RFLPs were also detected with MspI and PvuII, but the MspI RFLP correlated completely with CRC3 in this material and the PvuII RFLP could not be scored reliably due to a minute size difference between the two allelic fragments. The autosomal codominant inheritance of these RFLP loci was confirmed by family analyses. Significant evidence for genetic linkage between the CRC1/CRC3 loci and the A1BG locus in the HAL linkage group confirmed a previous assignment of the CRC gene to chromosome 6 in the pig.     

Novel detection of restriction fragment length polymorphisms in the human major histocompatibility complex

Cosmid genomic DNA clones have been used as hybridization probes in genomic Southern blot analysis to define restriction fragment length polymorphisms (RFLPs) in the major histocompatibility complex (MHC). Using 14 different enzymes and three overlapping cosmid clones we have detected six RFLPs in a 100 kilobase (kb) segment of DNA in the class III region extending centromeric of theTNFA gene towardHLA-DR. Four of the five RFLPs, defined using the enzymesTaqI,Rsa I,Hinc II, andHind III, and detected by the cosmid clone cosM7B, map to a 29 kb segment of DNA that includes all of the recently described G2 (BAT2) gene and a large portion of the 3 end of the G3 (BAT3) gene. The different RFLP variants were established by analyzing the DNA from three informative families and a panel of 51HLA-homozygous typing cell lines. CosM7B detectsTaq I variants of 4.3 kb, and 2.9 kb or 2.8 kb, Rsa I variants of 2.9 kb or 2.4 kb,Hinc II variants of 5.8 kb or 3.8 kb and 1.4 kb, and aHind III variant of 4.8 kb, while cosOT2 detects Taq I variants of 4.5 kb or 4 kb. The distribution of theRsa 1, Hinc II and Taq I RFLPs detected by cosM7B, and theTaq I RFLP detected with cosOT2, within the panel of cell line DNAs was assessed by Southern blotting. The 4.3 kbTaq I variant was observed in only one cell line with the extended haplotypeHLA-A29, C-, B44, SC30, DR4. The other RFLPs, however, occurred much more frequently. The 2.8 kb Taq I variant was observed in 20 % of haplotypes, the 2.9 kbRsa I variant was observed in 42% of haplotypes, and the 5.8 kbHinc I variant was observed in 12 % of haplotypes analyzed. The 4.5 kbTaq I variant detected by the overlapping cosmid cosOT2 was present in 21 % of haplotypes. Analysis of the RFLP variants with each other revealed seven different haplotypic combinations. Three of the haplotypic combinations were each subdivided into two subsets on the basis of the Nco I RFLP variant they carried at theTNF-B locus. These haplotypic combinations potentially allow differentiation among different extended haplotypes such asHLA-B8, SC01, DR3, HLA-B18, F1 C30, DR3, andHLA-B44, FC31, DR7. The RFLPs detected by the cosmid clones thus provide new tools which will be useful in the further genetic analysis of the MHC class III region.     

Polymorphic DNA haplotypes at the phenylalanine hydroxylase locus and their relation to phenotype in Swedish phenylketonuria families

Summary The genetic heterogeneity at the phenylalanine hydroxylase (PAH) locus was studied in 88 families including 93 of the 105 children with phenylketonuria (PKU) or hyperphenylalaninemia (HPA) detected through the Swedish neonatal screening program from 1966 to the end of 1986. Haplotypes based on eight restriction fragment length polymorphisms (RFLPs) at the PAH locus could be constructed for 132 normal and 136 mutant alleles. The normal alleles were of 27 different RFLP haplotypes, 9 of which have not been described previously, but there was a dominance of a few haplotypes common to many European populations. The distribution of mutant alleles was significantly different from that in neighboring countries, even though over 90% of all mutant alleles were confined to six RFLP haplotypes, also prevalent in other European populations. Allele-specific oligonucleotide hybridization analysis for the Arg⁴⁰⁸ to Trp⁴⁰⁸ mutation and for the G to A splicing mutation in intron 12 showed exceptions to the previously reported linkage of these mutations to mutant haplotypes 2 and 3, respectively. Correlation of mutant alleles with clinical phenotypes pointed to the presence of at least two different mutations associated with each of six haplotypes. We argue that PKU/HPA in the Swedish population may be caused by at least 13 different mutations in addition to the 4 already identified. The theoretical informativity of RFLP analysis in heterozygote detection and prenatal diagnosis in PKU/HPA families was estimated at approximately 85%. Carrier detection could, in effect, be accomplished for 88% of the 56 healthy siblings in the families studied.     

Diagnosis of genetic disease using recombinant DNA. Second edition

Summary Recombinant DNA methodology has greatly increased our knowledge of the molecular pathology of the human genome at the same time as providing the means to diagnose inherited disease at the DNA level. Direct detection and analysis of a range of genetic defects are now possible using cloned gene or oligonucleotide probes or by direct sequencing of the disease gene(s). In addition, the use of restriction fragment length polymorphism (RFLPs) within and around these genes as indirect genetic markers has now potentiated the tracking of disease alleles in affected pedigrees in cases where direct analysis was not feasible. RFLPs associated with linked anonymous segments may also be used not only to diagnose hitherto undetectable disease states, but also for chromosomal localization of the loci responsible. We present here an up-to date list of reports describing both the direct and the indirect analysis/diagnosis of human inherited disease, which is intended to serve as a guide to current molecular genetic approaches in diagnostic medicine.     

Genetic Analysis of a Mouse t Complex Locus That Is Homologous to a Kidney Cdna Clone

A mouse kidney cDNA clone, pMK174, identifies restriction fragment length polymorphisms (RFLPs) that map to two unlinked loci. One, designated D17Rp17, has been mapped near quaking, (qk), on chromosome 17 using three sets of recombinant inbred (RI) strains. A study of several t haplotypes resulted in the identification of t-specific alleles of D17Rp17 that map to the proximal half of the t complex. Neither t-specific nor wild-type D17Rp17 alleles are present in chromosomes carrying either the T Orleans (TtOrl) or the T hairpin tail (Thp) deletions. Comparison with other molecular markers indicates that pMK174 identifies a new proximal t complex locus, Rp17. The second locus identified by pMK174, termed D4Rp18, is tentatively assigned to chromosome 4 by mouse-Chinese hamster somatic cell hybrid analysis.     

Identification and characterization of nine RFLPs at the adenosine deaminase (ADA) locus.

We have identified and/or characterized at least nine RFLPs at the adenosine deaminase (ADA) locus, detected by digestion of DNA with MspI, BanII, PstI, BalI, and PvuII. The RFLPs were distributed over approximately 15 kb of the gene, from IVS 2 to IVS 10. They exhibited Mendelian inheritance and were in Hardy-Weinberg equilibrium. For seven fully characterized RFLPs, the gene frequencies of the rare alleles in 90 chromosomes examined ranged from .33 to .04, the PIC from .34 to .07, and the heterozygosity from .09 to .58. In kindreds examined (58 independent chromosomes), a total of nine haplotypes could be defined on the basis of seven fully characterized RFLPs with a heterozygosity of .62 and PIC of .53. Because there was considerable linkage disequilibrium, only three haplotypes accounted for 90% of individuals. Similar heterozygosity and PIC values (.59 and .51, respectively) could be obtained on the basis of haplotypes defined by the two sites that were the most polymorphic and that were in the least degree of linkage disequilibrium. A strategy for use of the RFLPs in linkage studies is suggested. We have also examined DNA from 17 patients with complete genetic deficiency of ADA (resulting in severe combined immunodeficiency [ADA-SCID] and from 10 patients with partial ADA deficiency (deficient in erythrocytes, with varying levels of ADA in other cells and normal immune function). Although the RFLPs detected genetic compounds among both types of patients, there was, as expected, a decreased incidence of heterozygosity (ADA-SCIDs, .29; partial ADA deficients, .20). Two additional haplotypes not found in the normal population were identified in homozygous form in patients. This information should be useful in developing a rational approach to delineation of mutations at the ADA locus as well as in distinguishing recurrent mutations of independent origin from those derived from a common progenitor.     

Macrophyte development in unimpacted lowland rivers in Poland

Three ecologically and morphologically distinct forms of Arctic charr (Salvelinus alpinus L.) have been identified in Loch Rannoch, Scotland, whose evolutionary status and origins are incompletely understood. A study was made of restriction fragment length polymorphism (RFLPs) detected variation in the D-loop, ND1 and cytochrome b regions of the mitochondrial genome, encompassing >3500 bp. Eight RFLP haplotypes were identified that clustered into three distinct clans based on restriction differences and into four clans based on sequence differences. Significant differences in RFLP frequencies were found among all morph groups. The pelagic morph was highly divergent from the two benthic forms, with the benthic forms having variants from only one genetic clan while the pelagic was dominated by a single variant from another clan. The relative divergence observed among benthic and pelagic forms is ~10 fold greater when nucleotide divergence among the haplotypes, as well as haplotype frequency differences, is taken into account. Sequence divergence between haplotypes in the two main clans is of a similar order to that between haplotypes in these clans and a charr from North America. In contrast, divergence among the two benthic morphs relates entirely to differences in haplotype frequencies. The study confirms the genetic distinctiveness of the pelagic and benthic forms as well as of the two benthic forms. It strongly supports previous evidence that the genetic divergence between the pelagic and benthic populations is allopatric in origin. Additionally, the results strongly suggest that the two benthic populations have undergone peripatric divergence through the sequential colonisation of the two basins by one lineage, followed by their spatial separation and reproductive isolation.     

Spatial analysis of nuclear and mitochondrial RFLP genotypes in populations of the chestnut blight fungus, Cryphonectria parasitica

Spatial structure of both nuclear and mitochondrial RFLPs were studied in several populations of the chestnut blight fungus, Cryphonectria parasitica, using a variety of spatial autocorrelation tests designed to detect nonrandom patterns. Fungal individuals were sampled from cankers on infected chestnut trees, and the location of each tree was mapped. Single-locus nuclear RFLPs, nuclear fingerprints, and mitochondrial DNA haplotypes were determined for each individual. Individuals with the same DNA fingerprint genotypes occurred closer together than would be expected at random in four of the five plots, while mitochondrial DNA haplotypes were aggregated in all five plots. Genetic distances between individuals, expressed as one minus the proportion of shared restriction fragment size classes for fingerprints and mitochondrial haplotypes, were significantly correlated with Euclidean distances between individuals in four of the five populations, but these correlations were very weak (r < 0.18). The same DNA fingerprint and single-copy nuclear RFLP alleles occurred on the same trees or immediately neighbouring trees more often than would be expected at random. Most of the aggregation for all three genetic markers occurred among individuals within the same cluster of chestnut stems or on neighbouring trees. Lack of spatial autocorrelation in one population was probably due to sampling on a larger scale that was too coarse to detect any patterns. Significant aggregation of genotypes in C. parasitica is most likely caused by some degree of restricted dispersal within populations. The implications of restricted dispersal are discussed in relation to the breeding system and isolation by distance in populations of. C. parasitica.     

Nuclear and chloroplast DNA differentiation in Andean potatoes.

Over 3500 accessions of Andean landraces have been known in potato, classified into 7 cultivated species ranging from 2x to 5x (Hawkes 1990). Chloroplast DNA (ctDNA), distinguished into T, W, C, S, and A types, showed extensive overlaps in their frequencies among cultivated species and between cultivated and putative ancestral wild species. In this study, 76 accessions of cultivated and 19 accessions of wild species were evaluated for ctDNA types and examined by ctDNA high-resolution markers (ctDNA microsatellites and H3 marker) and nuclear DNA restriction fragment length polymorphisms (RFLPs). ctDNA high-resolution markers identified 25 different ctDNA haplotypes. The S- and A-type ctDNAs were discriminated as unique haplotypes from 12 haplotypes having C-type ctDNA and T-type ctDNA from 10 haplotypes having W-type ctDNA. Differences among ctDNA types were strongly correlated with those of ctDNA high-resolution markers (r = 0.822). Differentiation between W-type ctDNA and C-, S-, and A-type ctDNAs was supported by nDNA RFLPs in most species except for those of recent or immediate hybrid origin. However, differentiation among C-, S-, and A-type ctDNAs was not clearly supported by nDNA RFLPs, suggesting that frequent genetic exchange occurred among them and (or) they shared the same gene pool owing to common ancestry.     

Molecular definition of red cell Rh haplotypes by tightly linked SphI RFLPs.

The Rh blood group system of human red cells contains five major antigens D, C/c, and E/e (the latter four designated "non-D") that are specified by eight gene complexes known as Rh haplotypes. In this paper, we report on the mapping of RH locus and identification of a set of SphI RFLPs that are tightly linked with the Rh structural genes. Using exon-specific probes, we have localized the SphI cleavage sites resulting in these DNA markers and derived a comprehensive map for the RH locus. It was found that the SphI fragments encompassing exons 4-7 of the Rh genes occur in four banding patterns or frameworks that correspond to the distribution and segregation of the common Rh haplotypes. This linkage disequilibrium allowed a genotype-phenotype correlation and direct determination of Rh zygosity related to the Rh-positive or Rh-negative status (D/D, D/d, and d/d). Studies on the occurrence of SphI RFLPs in a number of rare Rh variants indicated that Rh phenotypic diversity has taken place on different haplotype backgrounds and has arisen by diverse genetic mechanisms. The molecular definition of Rh haplotypes by SphI RFLP frameworks should provide a useful procedure for genetic counseling and prenatal assessment of Rh alloimmunization.     

Gene duplications and sequence polymorphism of bovine class II DQB genes

The genetic diversity of bovine class II DQB genes was investigated by polymerase chain reaction amplification and DNA sequencing. The first domain exon was amplified from genomic DNA samples representing 14 class II haplotypes, defined by restriction fragment length polymorphism (RFLP) analysis. The presence of a polymorphism in the copy number of DQB genes was confirmed since two DQB sequences were isolated from certain haplotypes. Four subtypes of bovine DQB genes were found. DQB1 is the major type and was found in almost all haplotypes. DQB2 is very similar DQB1 but was found only in the duplicated haplotypes DQ9 to 12. DQB3 and DQB4 are two quite divergent genes only present in certain duplicated haplotypes. The bovine DQB complexity thus resembles that in the human DRB region. Bovine DQB genes were found to be highly polymorphic as ten DQB1 alleles and four DQB2 alleles were identified. The observed sequence polymorphism correlated well with previously defined DQB RFLPs. Bovine and human DQB alleles show striking similarities at the amino acid level. In contrast, the frequency of silent substitutions is much higher in comparisons of DQB alleles between species than within species ruling out the possibility that any of the contemporary DQB alleles have been maintained since the divergence of humans and cattle. The frequency of silent substitutions between DQB alleles was markedly lower in cattle than in humans, in agreement with a previous comparison of human and bovine DRB alleles.     

Diagnosis of human genetic disease using recombinant DNA

Recombinant DNA methodology has greatly increased our knowledge of the molecular pathology of the human genome at the same time as providing the means of diagnosing inherited disease at the DNA level. Direct detection and analysis of a wide range of genetic lesions are now possible using cloned gene or oligonucleotide probes or by direct sequencing of the disease gene(s). In addition, the use of restriction fragment length polymorphisms (RFLPs) within and around these genes as indirect genetic markers has potentiated the tracking of disease alleles in affected pedigrees in cases where direct analysis is not yet feasible. RFLPs associated with linked anonymous DNA segments may also be used not only to diagnose hitherto undetectable disease states, but also for the chromosomal localization of the loci responsible. We present here an update to our previous list of reports describing the direct and indirect analysis/diagnosis of human inherited disease. This compilation is intended to serve as a guide to current molecular genetic approaches in diagnostic medicine.     

The use of restriction fragment length polymorphisms in paternity analysis.

This paper examines the utility of restriction fragment length polymorphisms (RFLPs) for paternity analysis. While, on the average, 99% of falsely accused males can be excluded with the standard battery of blood group antigens, red cell enzymes, serum proteins, and HLA antigens, there are still mother-child pairs for whom the exclusion probability is not high. It has been suggested that additional resolution would be available with RFLPs. We have examined the strategic aspects of using RFLPs for paternity analysis, comparing the efficacy and cost of a multimarker haplotypic set with those of a comparable set of unlinked RFLPs, using published frequencies for the beta-globin complex, the serum albumin region, and the growth hormone region. There are four major findings. (1) Greater resolution is obtained with a carefully chosen set of tightly linked RFLPs producing chromosomal haplotypes than with a comparable set (same allele frequencies) of unlinked markers, but only if it is possible to establish linkage phase unambiguously. (2) Assay of linked sets is cheaper than is the assay of unlinked markers, but the cost advantage is optimized with sets of no more than two or three linked markers. (3) Also, with more than two or three tightly linked markers, the haplotypic frequencies are too poorly estimated to provide a reliable measure of the probability of paternity for unexcluded males, given the sample sizes likely to be available in the near future. (4) Optimal resolution, minimal cost, and acceptable accuracy are obtained with several independent sets of no more than two or three tightly linked RFLP markers each. With current technology, RFLP analysis is more expensive for the same level of genetic resolution than is the standard battery, but gradual replacement of the latter can be anticipated as economies of scale reduce the cost of the DNA technology.     

Genetic diversity among progenitors and elite lines from the Iowa Stiff Stalk Synthetic (BSSS) maize population: comparison of allozyme and RFLP data

Summary Data for restriction fragment length polymorphisms (RFLPs) of 144 clone-enzyme combinations and for 22 allozyme loci from 21 U.S. Corn Belt maize (Zea mays L.) inbreds were analyzed. The genetic materials included 14 progenitors of the Iowa Stiff Stalk Synthetic (BSSS) maize population, both parents of one missing BSSS progenitor, four elite inbreds derived from BSSS, and inbred Mo17. Objectives were to characterize the genetic variation among these 21 inbreds for both allozymes and RFLPs, to compare the results from both types of molecular markers, and to estimate the proportion of unique alleles in the BSSS progenitors. Genetic diversity among the 21 inbreds was substantially greater for RFLPs than for allozymes, but the percentages of unique RFLP variants (27%) and unique allozyme alleles (25%) in the BSSS progenitors were similar. Genetic distances between inbreds, estimated as Rogers' distance (RD), were, on average, twice as large for RFLP (0.51) as for allozyme data (0.24). RDs obtained from allozyme and RFLP data for individual line combinations were only poorly correlated (r = 0.23); possible reasons for discrepancies are discussed. Principal component analysis of RFLP data, in contrast to allozyme data, resulted in separate groupings of the ten BSSS progenitors derived from the Reid Yellow Dent population, the four BSSS elite lines, and Mo17. The remaining six BSSS progenitors were genetically rather diverse and contributed a large number of rare alleles to BSSS. The results of this study corroborate the fact that RFLPs are superior to allozymes for characterizing the genetic diversity of maize breeding materials, because of (1) the almost unlimited number of markers available and (2) the greater amount of polymorphisms found. In particular, RFLPs allow related lines and inbreds with common genetic background to be identified, but a large number of probe-enzyme combinations is needed to estimate genetic distances with the precision required.Joint contribution from Cereal and Soybean Research Unit, USDA, Agricultural Research Service, and Journal Paper No. J-14236 of the Iowa Agricultural and Home Economics Experiment Station, Projects 2818 and 2778     

The Putative Oncogene Pim-1 in the Mouse: Its Linkage and Variation among t Haplotypes

Pim-1, a putative oncogene involved in T-cell lymphomagenesis, was mapped between the pseudo-alpha globin gene Hba-4ps and the alpha-crystallin gene Crya-1 on mouse chromosome 17 and therefore within the t complex. Pim-1 restriction fragment variants were identified among t haplotypes. Analysis of restriction fragment sizes obtained with 12 endonucleases demonstrated that the Pim-1 genes in some t haplotypes were indistinguishable from the sizes for the Pim-1b allele in BALB/c inbred mice. There are now three genes, Pim-1, Crya-1 and H-2 I-E, that vary among independently derived t haplotypes and that have indistinguishable alleles in t haplotypes and inbred strains. These genes are closely linked within the distal inversion of the t complex. Because it is unlikely that these variants arose independently in t haplotypes and their wild-type homologues, we propose that an exchange of chromosomal segments, probably through double crossingover, was responsible for indistinguishable Pim-1 genes shared by certain t haplotypes and their wild-type homologues. There was, however, no apparent association between variant alleles of these three genes among t haplotypes as would be expected if a single exchange introduced these alleles into t haplotypes. If these variant alleles can be shown to be identical to the wild-type allele, then lack of association suggests that multiple exchanges have occurred during the evolution of the t complex.     

Restriction fragment length polymorphism analysis of the kappa-casein locus in cattle

The two common genetic variants (A and B) of bovine kappa-casein originate from two point mutations in the codons for the aminoacids in position 136 and 148. These mutations give rise to polymorphic sites for the restriction endonucleases Hin dIII, AluI, HinfI, Mbo II and TaqI. We have examined DNAs of several Italian Friesian cows and bulls of known and unknown genotype by Southern analyses using kappa-casein cDNA probes. Restriction fragment length polymorphisms (RFLPs) specific for the A and B alleles were identified for each of the above enzymes, except for AluI, which has a non-polymorphic site 12bp away from the polymorphic one. We have also found two new polymorphic sites for MboII and TaqI in the non-coding regions. These sites differentiate the A allele into two new variants, named A1 and A2. The RFLP analysis permits the characterization of kappa-casein alleles even in the absence of their expression. This should facilitate selective breeding programmes aimed at increasing the frequency of the kappa-casein B allele whose product improves the cheesemaking properties of milk.     

Restriction fragment length polymorphisms of the angiotensinogen gene in inbred rat strains and mapping of the gene on chromosome 19q

Using a cDNA probe of the rat angiotensinogen gene (ANG), restriction fragment length polymorphisms (RFLPs) were detected in inbred rat strains with the restriction enzymes HindIII, PstI, and PvuII. Three alleles of ANG were almost equally distributed in 11 inbred strains. In two sets of backcross progeny originating from parental strains with different alleles, no close linkage was found between the ANG locus and 17 other loci tested. In situ hybridization, however, allowed assignment of the gene to chromosome 19q. The RFLPs of the angiotensinogen gene, therefore, can be considered useful as markers of rat chromosome 19.