Restriction Fragment Length Polymorphism Genotyping Research Articles

Effects of Heart Fatty Acid-binding Protein Genotype on Intramuscular Fat Content in Duroc Pigs Selected for Meat Production and Meat Quality Traits

Using multi-trait animal model BLUP, selection was conducted over seven generations for growth rate (DG), real-time ultrasound loin-eye muscle area (LEA), backfat thickness (BF), and intramuscular fat content (IMF) to develop a new line of purebred Duroc pigs with enhanced meat production and meat quality. This study was intended to investigate the relationship between restriction fragment length polymorphism (RFLP) of a heart fatty acid-binding protein (H-FABP) gene and intramuscular fat content (IMF) of this Duroc purebred population. The present experiment examined the RFLP of 499 slaughtered pigs. The DNA was separated from the blood or ear tissue of the pigs, which were slaughtered at 105 kg of body weight. Intramuscular fat content of the longissimus muscle was measured using chemical analysis. A significant difference was detected in the breeding value of IMF among the H-FABP PCR RFLP genotypes. The AA genotype has a significantly larger positive effect on the IMF breeding value than do the Aa and aa genotypes for the MspI RFLP. In addition, the DD genotype has a significantly greater positive effect on IMF breeding value than the Dd and dd genotypes for the HaeIII RFLP. For the HinfI RFLP, the hh genotype has a significantly larger positive effect on IMF breeding value than the HH genotype. Multiple regression analysis was performed using the IMF breeding values as the dependent variable and the three H-FABP genotypes as independent variables. Results revealed that the contribution of the genotypes to variation in IMF breeding values was approximately 40%. These results demonstrated that H-FABP RFLPs affect IMF in this Duroc population.

Angiotensin I‐converting enzyme insertion‐related genotypes and allele are associated with higher susceptibility of endometriosis and leiomyoma

Endometriosis and leiomyoma display features similar to malignancy, requiring neovascularization to proliferation and growth. Altered vascular-related genes might be related to the development of endometriosis and leiomyoma. Polymorphisms of the angiotensin-converting enzyme (ACE) insertion/deletion (I/D) genes have been linked with some vascular diseases. This study investigates whether ACE I/D gene polymorphisms could be used as markers of susceptibility in endometriosis and leiomyoma. Women were divided into three groups: (1) endometriosis (n = 125); (2) leiomyoma (n = 120); (3) normal controls (n = 128). Genomic DNA was obtained from peripheral leukocyte. ACE I/D gene polymorphisms in intron 16 were amplified by polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP) Genotypes and allelic frequencies in both groups were compared. We observed the genotype distribution and allele frequency of ACE I/D gene polymorphisms in both groups were significantly different. Proportions of ACE*I homozygote/heterozygote/D homozygote in both groups were: (1) 50.4/24/25.6%; (2) 25/23.33/51.67%; (3) 10.2/29.7/60.1%. Proportions of I/D alleles in each group were: (1) 62.4/37.6%; (2) 36.7/63.3%; (3) 25/75%. We concluded that ACE*I/D gene polymorphisms are associated with endometriosis and leiomyoma susceptibilities. ACE*I-related genotypes and allele are strongly related to the occurrence of endometriosis and moderately related to the occurrence of leiomyoma.

Genetic Diversity of Naturally Established Ectomycorrhizal Fungi on Norway Spruce Seedlings under Nursery Conditions

We have assessed ectomycorrhizal fungi colonizing Norway spruce (Picea abies L.) seedlings in nine forest nurseries using restriction fragment length polymorphism (RFLP) and sequencing analyses of the internal transcribed spacers (ITS1-5.8S-ITS2) amplicons. Restriction analysis of the amplified DNA fragments with HinfI, MboI, and TaqI enzymes allowed the definition of 17 RFLP genotypes; five of them could be unambiguously assigned to Thelephora terrestris, Hebeloma longicaudum, H. crustuliniforme, Tricharina ochroleuca, and Cenococcum geophilum species by comparison with the sporocarp RFLP-pattern database. The remaining genotypes have been sequenced and compared with sequences deposited in the GenBank database. The phylogenetic analysis of resulting sequences and their identified matches indicated that isolated genotypes have formed seven clades. The ascomycetes were predominant: we have determined eight species--Wilcoxina mikolae, Phialophora finlandia, Tuber sp., Cenococcum geophilum, Tricharina ochroleuca, Pulvinula constellatio, and two unidentified ascomycetes--whereas the basidiomycetes were less common (four species denoted: Amphinema byssoides, Hebeloma crustuliniforme, H. longicaudum, and Thelephora terrestris). Wilcoxina mikolae and Phialophora finlandia were the most frequent fungi. Analysis of variance revealed that ascomycetes abundance was higher in nurseries that used organic fertilizer.

Identification of a Novel C5L2 Variant (S323I) in a French Canadian Family With Familial Combined Hyperlipemia

A functional acylation stimulating protein (ASP) receptor, C5L2, has been recently identified in ASP-responsive cells. Impaired ASP-mediated triglyceride synthesis has previously been described in a subset of hyperapolipoprotein B/familial combined hyperlipidemia subjects. DNA sequencing of C5L2 coding region in 61 unrelated probands identified a heterozygous variant (G968-->T) in 1 subject, resulting in Ser323-->Ile substitution in the carboxyl terminal region. This variant was not detected in 2176 additional chromosomes by restriction fragment length polymorphism or fluorescence polarization genotyping. Eight family members of the proband were identified with one altered (+/-)C5L2 allele. Nine other family members had the wild-type (+/+)C5L2 sequence. The abnormal allele was associated with increased plasma triglyceride, plasma cholesterol, low-density lipoprotein (LDL) cholesterol, apolipoprotein B and ASP. Of 23 subjects tested in cell-based ASP bioactivity assays, those with C5L2(+/-) variant (n = 2) had a 50% reduction in ASP-stimulated triglyceride synthesis, glucose transport and marked reduction in maximal binding (B(max)). By contrast, a C5L2(+/+) family member responded normally, as did hyperapolipoprotein B normal ASP subjects compared with C5L2(+/+) controls (n = 6). The S323I variant may alter C5L2 function and might be one molecular basis contributing to familial combined hyperlipidemia.

High-throughput genotyping with infrared fluorescence allele specific hybridization (iFLASH): A simple, reliable and low-cost alternative

Objectives To develop and validate a novel genotyping approach, named infrared Fluorescence Allele Specific Hybridization (iFLASH), which combines the principles of allele specific oligonucleotide (ASO) hybridization with the advanced possibilities of infrared imaging. Design and methods As an example, we genotyped the 55L > M and the 192Q > R common genetic variants of the paraoxonase-1 gene in 92 DNA samples using the iFLASH technique, and validated the outcomes with the restriction fragment length polymorphism (RFLP) and TAQman genotyping assays. Results There was a 100 percent agreement in genotype outcome among the three methods. Conclusions Although we found complete unity in genotype outcome, the iFLASH assay has essential advantages over the RFLP and TAQman genotyping assays. First, the iFLASH technique is capable of handling up to 1536 samples per assay, which makes it a suitable technique for high-throughput genotyping. Secondly, because the costs per assay are lower, high-throughput genotyping with iFLASH is affordable.

Internal Restriction Sites: Quality Assurance Aids in Genotyping

Improvements to restriction fragment length polymorphism (RFLP)-based genotyping assays currently used for detection of mutations responsible for bovine ferrochelatase and myophosphorylase deficiencies, and equine hyperkalemic periodic paralysis (HYPP) are described. Reports of sporadic inhibition of restriction enzyme activity suggest a critical factor in RFLP-based genotyping assays should be assurance that restriction enzymes perform to specification with every sample. The RFLP genotyping assays that use either a mismatched recognition sequence in one or both of the oligonucleotides, or incorporate a second native site within the PCR amplicon, provide the mechanism by which efficiency of restriction enzymes can be assessed with every sample. The outcome is confirmation of the activity of the discriminating enzyme regardless of genotype.

Isolation of Enterobacter sakazakii from Stable Flies, Stomoxys calcitrans L. (Diptera: Muscidae)

Enterobacter sakazakii is an opportunistic foodborne pathogen that causes meningitis, enterocolitis, and sepsis, primarily in immunocompromised infants. Previously, it was suggested that stable flies, Stomoxys calcitrans, were a vector or reservoir of this pathogen. In our study, by means of a culturing approach combined with 16S rDNA PCR–restriction fragment length polymorphism genotyping and sequencing, we screened 928 individual stable flies collected in Kansas and Florida. Two stable flies (0.2%) were positive for E. sakazakii. In addition, 411 (44%) stable flies carried bacteria-forming red colonies (presumably enterics) on a violet red bile glucose agar (mean count = 6.4 × 104 CFU per fly), and 120 (13%) stable flies carried fecal coliforms (mean count = 8.7 × 103 CFU per fly). Sequencing of 16S rDNA showed that enterics from violet red bile glucose agar were represented by several genera, including Escherichia, Shigella, Providencia, Enterobacter, Pantoea, Proteus, Serratia, and Morganella. Our study shows that stable flies carry bacteria typically present in animal manure (a developmental site of stable fly larvae), which indicates that the natural reservoir of E. sakazakii is the digestive tract or manure of domestic animals. The low prevalence of E. sakazakii associated with stable flies suggests that stable flies do not play a major role as a reservoir or vector of this pathogen.

Genetic diversity in fluoroquinolone and macrolide-resistant Campylobacter coli from pigs

The genetic diversity of 115 Campylobacter coli strains, isolated from pigs of 59 geographical distant farms in Switzerland, were characterized on the basis of their DNA fingerprints and resistance to macrolides and fluoroquinolones. Sequence analysis showed that the macrolide-resistant isolates had a point mutation in the 23S ribosomal RNA (rRNA) genes (A2075G) and that the fluoroquinolone-resistant isolates had a point mutation in the gyrase gene gyrA (C257T). One fluoroquinolone-resistant strain had an additional transition mutation in the gyrB gene (A1471C). The flaA restriction fragment length polymorphism (RFLP) genotyping revealed that 57% of the isolates were genetically different. Point mutations in the 23S rRNA and gyrA genes could be found in both genetically distant and genetically related isolates. Additionally, isolates with and without point mutations were found within individual farms and on different farms. This study showed that the ciprofloxacin and erythromycin-resistant C. coli population present on the pig farms is not issued from a common ancestral clone, but individual Campylobacter strains have most likely mutated independently to acquire resistances under the selective pressure of an antibiotic.

Comparison of disseminated and nondisseminated strains of Borrelia burgdorferi sensu stricto in mice naturally infected by tick bite.

Clinical isolates of Borrelia burgdorferi sensu stricto have been categorized into disseminated and nondisseminated groups based on distinct ribosomal spacer restriction fragment length polymorphism genotypes (RSTs). In order to determine whether transmission by tick bite would alter the dissemination dynamics and disease produced by distinct genotypes, disseminated isolates (RST1), nondisseminated isolates (RST3), and a standard laboratory strain (B-31) were established in a murine cycle utilizing infections transmitted by ticks. B-31 spirochetes circulated in the blood of inbred C3H/HeJ mice longer than in the blood of outbred mice. The majority of C3H mice exposed to RST1-infected ticks contained cultivable spirochetes in their blood for up to 17 days; in contrast, mice exposed to RST3 isolates demonstrated a precipitous decline in infection after day 7 postexposure. A quantitative PCR (q-PCR) assay demonstrated that the densities of spirochetes in blood were similar for the RST1 and RST3 isolates, except during the 2nd week postexposure, when the RST1 isolates displayed a markedly higher density in blood. Spirochete load in the heart and bladder of infected mice was measured by q-PCR at 8 weeks postexposure; the numbers of spirochetes in these tissues were similar for mice infected with either disseminated or nondisseminated strains. Similarly, histopathology samples of heart, bladder, and joint tissue obtained at 8 weeks postexposure did not reveal greater pathology in mice infected with the disseminated isolates. We conclude that although the spirochetemia induced by tick-transmitted disseminated isolates was more intense and of longer duration than that induced by nondisseminated isolates, the resultant pathologies produced by these strains were ultimately similar.

Lack of association of the vitamin D receptor gene with Graves' disease in UK Caucasians.

Vitamin D modulates the immune system by suppressing the proliferation of activated T cells, with its actions being directed through the vitamin D receptor (VDR). A number of single nucleotide polymorphisms (SNPs) have been identified in the VDR gene, of which several have been associated with autoimmune diseases, including type 1 diabetes and Graves' disease (GD) in Japanese females. The aim of this study was to test for association of polymorphisms of the VDR gene in the genetic susceptibility to GD in UK Caucasians. Target DNA for five previously published SNPs, four novel SNPs and one microsatellite marker was amplified by the polymerase chain reaction (PCR). Subsequent genotyping was performed using restriction fragment length polymorphism (RFLP) or microsatellite genotyping analysis, according to the type of VDR polymorphism. We obtained DNA from a case-control dataset consisting of 768 patients with GD and 864 control subjects. All patients and control subjects were Caucasians born in the UK, and all gave informed, written consent. Frequencies of the alleles and genotypes of the ten VDR gene polymorphisms were compared between patients and control subjects using the chi2 test. Odds ratios were calculated using Woolf's method with Haldane's modification for small numbers and D prime (D') was calculated to assess the level of linkage disequilibrium (LD) between the ten polymorphisms. No differences in allele or genotype frequencies were observed between GD cases and control subjects for any of the nine SNPs studied. The S allele of the PolyA microsatellite marker was slightly more frequent in GD cases when compared with control subjects (chi2= 4.364, P = 0.04). Strongest LD between markers was observed towards the 3' end of the VDR gene but there was no evidence of association with disease. This is the largest and most comprehensive study of the VDR gene in GD to date and these data suggest that these polymorphisms of the VDR gene do not contribute to GD susceptibility in the UK.

Development of a multigenic oligoligation assay test in relation to meat and carcass traits in cattle

SummarySimultaneous with the increased interest in the use of single nucleotide polymorphisms (SNPs) in genetic studies (such as association studies), the demand for high‐throughput SNP genotyping has risen. In order to accommodate the high throughput SNP analysis, the oligoligation assay (OLA) technique was adapted to genotype simultaneously 11 mutations in seven different candidate genes, related to meat and carcass traits in cattle. The accuracy of the multigenic test was determined by comparing the OLA data with previously obtained restriction fragment length polymorphism (RFLP) genotypes. Both data sets were 100% identical. The accuracy of the multigenic test coupled with the high throughput, relatively low cost (once optimized) and simplicity of test ensures that this test can be routinely used in each laboratory. Furthermore, the flexibility of this test means that genes can be replaced or that new ones can be added. Although initially this multigenic test was designed as a basic tool for association studies, it can be easily used as a diagnostic tool. The obtained genotype information can be integrated in breeding schemes in order to genetically enhance the herd.

Genetic differences in wild strains of Lentinula edodes collected from a single fallen tree

Genetic differences among 18 Lentinula edodes strains isolated from a fallen trunk of Quercus mongolica var. grosseserrata were characterized by mating tests and restriction fragment length polymorphism (RFLP) analyses of mitochondrial DNA (mtDNA). These strains could be divided into six genets of different mating types. Because the mtDNA of the 18 strains showed four different RFLP genotypes, these strains seemed to have originated from at least 4 distinct parental strains. Strains belonging to the same genet were collected from fruiting bodies located not more than about 1m apart on the fallen tree. Implications of these findings regarding the degree of genetic variation and territory sizes of individual genets of wood-decaying basidiomycetes such as L. edodes are discussed.

Glycoprotein B genotyping of cytomegalovirus strains isolated in a pediatric population

To better characterize cytomegalovirus (CMV) isolates in a pediatric population, we performed glycoprotein B genotyping of isolates from CMV culture-positive patients during a 3-year period at Children's Hospital Medical Center, Cincinnati, Ohio. The most common sources for CMV isolation were respiratory and urine cultures. Among 61 isolates glycoprotein B genotype distribution was genotype 1, 32%; genotype 2, 24%; genotype 3, 32%; and genotype 4, 10%. Among viremic isolates there was a trend favoring type 1 genotype (P = 0.057).

Candidate genes for growth traits in beef cattle crosses Bos taurus × Bos indicus

SummaryCandidate gene strategy has been proposed to direct the search for quantitative trait loci, and polymorphisms at these loci have been associated with several production traits in bovine. A total of 211 animals resulting from crosses between Nelore females with sires from either Aberdeen Angus, Canchim and Simmental breeds were analyzed for restriction fragment length polymorphisms (RFLPs) in the κ‐casein, β‐lactoglobulin (LGB) and growth hormone GH genes. Average daily gain from birth to weaning (GBW) and from weaning to yearling (GWY) were analyzed by a statistical model comprising RFLP genotypes and their interaction effects. Significant effects (p < 0.05) of growth hormone (GH) and of the interaction β‐lactoglobulin × GH were verified for the GWY. Animals with LL genotypes for GH had higher GBW than LV animals but lower GWY (p < 0.05).

Genetic characterization of Aberdeen Angus cattle using molecular markers

Aberdeen Angus beef cattle from the Brazilian herd were studied genetically using restriction fragment length polymorphism (RFLP) of the kappa-casein - HinfI (CSN3 - HinfI), beta-lactoglobulin - HaeIII (LGB - HaeIII) and growth hormone AluI (GH- AluI) genes, as well as four microsatellites (TEXAN15, CSFM50, BM1224 and BM7160). The RFLP genotypes were determined using the polymerase chain reaction (PCR) followed by digestion with restriction endonucleases and electrophoresis in agarose gels. With the exception of the microsatellite BM7160, which was analyzed in an automatic sequencer, the PCR products were genotyped by silver staining. The allele and genotype frequencies, heterozygosities and gene diversity were estimated. The values for these parameters of variability were comparable to other cattle breeds. The genetic relationship of the Aberdeen Angus to other breeds (Caracu, Canchim, Charolais, Guzerath, Gyr, Nelore, Santa Gertrudis and Simmental) was investigated using Nei's genetic distance. Cluster analysis placed the Aberdeen Angus in an isolated group in the Bos taurus breeds branch. This fact is in agreement with the geographic origin of this breed.

Erythrocyte CR1 expression level does not correlate with a HindIII restriction fragment length polymorphism in Africans; implications for studies on malaria susceptibility.

Complement receptor 1 (CR1) expression level on erythrocytes is genetically determined, and in Caucasian populations is linked to high (H) and low (L) expression alleles identified by a HindIII restriction fragment length polymorphism (RFLP). Erythrocyte CR1 may be an important factor in determining malaria susceptibility, as low expression of CR1 reduces the rosetting of uninfected erythrocytes with Plasmodium falciparum-infected cells, a process that contributes to malaria pathogenesis. Prior to studying CR1 expression and malaria susceptibility, we have investigated whether the quantity of erythrocyte CR1 correlates with the H and L alleles in an African population. Mean erythrocyte CR1 in 149 Malian adults was 415 molecules per cell, which is comparable to Caucasian populations; however, there was no relationship between erythrocyte CR1 level and genotype for the HindIII RFLP (mean CR1 per erythrocyte HH = 414, HL = 419 and LL = 403, P > 0.1, Student's t-test). The conclusions of a previous study of erythrocyte CR1 expression level and malaria susceptibility in West Africa that was based on HindIII RFLP genotyping may therefore need to be re-evaluated.

Disease severity in a murine model of lyme borreliosis is associated with the genotype of the infecting Borrelia burgdorferi sensu stricto strain.

The pathogenicity of Borrelia burgdorferi sensu stricto clinical isolates representing 2 distinct ribosomal DNA spacer restriction fragment-length polymorphism genotypes (RSTs) was assessed in a murine model of Lyme disease. B. burgdorferi was recovered from 71.5% and 26.6% of specimens from mice infected with RST1 and RST3 isolates, respectively (P<.0001). The average ankle diameter and histologic scores for carditis and arthritis were significantly higher after 2 weeks of infection among mice infected with RST1 isolates than among those infected with RST3 isolates (P<.001). These clinical manifestations were associated with larger numbers of spirochetes in target tissues but not with the serum sensitivity of the individual isolates. Thus, the development and severity of disease in genetically identical susceptible hosts is determined mainly by the pathogenic properties of the infecting B. burgdorferi isolate. The RST1 genotype is genetically homogeneous and thus may represent a recently evolved clonal lineage that is highly pathogenic in humans and animals.

Polymorphism of the vitamin D3 receptor in patients with psoriasis

1,25-Dihydroxyvitamin D is the biologically active form of vitamin D for the treatment of skin eruptions in patients with psoriasis. 1,25-(OH)(2)D(3) elicits its action on skin eruptions through the vitamin D receptor (VDR). Allelic frequencies of VDR were studied in 86 normal subjects and 50 patients with psoriasis. Genomic DNA was extracted from peripheral blood leukocytes and the VDR gene was amplified using a heminested polymerase chain reaction (PCR). The products were digested with respective restriction enzymes ApaI, TaqI and BsmI. The restriction fragment length polymorphisms (RFLP) were coded as Aa, Tt or Bb. The frequencies of ApaI, BsmI and TaqI RFLP genotypes in psoriasis patients showed no significant differences compared with normal controls. The frequency of the AA genotype was significantly higher in pustulosis palmaris et plantaris patients than in psoriasis vulgaris patients ( P<0.05), and in psoriasis vulgaris patients than in psoriasis pustulosa patients ( P<0.01). In patients with psoriasis, the levels of serum alanine 2-oxoglutarate aminotransferase (ALT) were significantly higher in patients with the AA genotype (54.0+/-22.0 IU/l, n=4) than in those with the aa genotype (24.0+/-15.9 IU/l, n=27; P<0.02). The distribution of ApaI, BsmI, TaqI RFLP VDR genotypes showed no significant relationship to the PASI score, serum aspartate 2-oxoglutarate aminotransferase or triglyceride levels, or age at onset. These results show that the VDR genotype contributes to the liver dysfunction in patients with psoriasis, although no correlation was found between VDR genotype and the skin eruptions of psoriasis.

Molecular typing of Neisseria gonorrhoeae from hospital and community isolates by restriction fragment length polymorphism.

Strains of Neisseria gonorrhoeae are generally characterized by auxotyping, serotyping, plasmid profile, antibiotic sensitivity and deoxyribonucleic acid (DNA) amplification fingerprinting. The aim of this study was to analyse the generation of restriction fragment length polymorphism (RFLP) patterns by BgIII digestion of total genomic DNA of N. gonorrhoeae isolated from the community (n =30) and the hospital (n =15) and to establish an association with serogrouping and antibiogram. The RFLP patterns produced by BgIII restriction digestion showed 7 different patterns among 30 community isolates and 9 different patterns among 15 hospital isolates. 66.7% of isolates belonged to serogroup WI. Penicillin resistance was observed in 46.7% of community isolates and 66.7% hospital isolates. However, penicillinase producing N. gonorrhoeae (PPNG) were lower in the community (6.6%) than in the hospital isolates (53.3%). PPNG strains were more often seen in serogroup WI. This is the first Indian report on RFLP genotype pattern in N. gonorrhoeae. We noted differences in RFLP genotypes of the community (RFLP types 1, 2, 3, 4, 5 and 7) and hospital strains (RFLP types 6 and 8), while no differences in the serogroup were observed. Ciprofloxacin resistance was 20.0% and 26.6% in the community and hospital isolates, respectively. Ceftriaxone emerges as the current drug of choice for an effective policy of antibiotic treatment of gonorrhoea through syndromic management in developing countries.

Microsomal epoxide hydrolase polymorphism and susceptibility to ovarian cancer

Polymorphic variants of microsomal epoxide hydrolase (mEPHX) with altered enzyme activity have been associated with an increased risk for ovarian cancer. We assessed the frequency of exon 3 and exon 4 variants of mEPHX among 291 ovarian cancers and 257 controls from a UK-based population. The distribution of the exon 3 alleles among both the cancer and control groups was significantly different from that expected under Hardy–Weinberg equilibrium suggesting that the PCR restriction fragment length polymorphism (PCR-RFLP) genotyping assay might be flawed. The codon 113 polymorphism was reassessed using a two-color allele-specific PCR-based assay. We found that a codon 119 G>A polymorphism, present in 20% of the British population and linked to the wild-type exon 3 allele, resulted in some Tyr113/His113 heterozygotes being falsely classified as His113/His113 homozygotes when using the PCR-RFLP assay. Consequently, we reassessed all our codon 113 data using the new allele-specific assay. We found no evidence of an association of ovarian cancer risk with the exon 3 Tyr113>His113 variant. Similarly the frequencies of the exon 4 His139>Arg139 genotypes were not significantly different between cases and controls. Stratifying the genotyping data according to the predicted mEPHX activity revealed a highly significant decrease in high mEPHX activity among the serous ovarian cancers ( P=0.01) suggesting that high mEPHX activity may be protective for this histological sub-type. Furthermore previous disease association studies of exon 3 alleles which utilized the PCR-RFLP assay may be compromised by the existence of a codon 119 G>A polymorphism which may be common in Caucasian populations.