Fragment Of Exon Research Articles

Hydrophobic core formation in protein complex of cathepsin

The “fuzzy oil drop” model assumes that the idealized hydrophobic core in a protein body can be described by a 3D Gauss function. The structure of the 1ICF protein (cathepsin), which participates in the proteolysis process and has cysteine-type peptidase activity, has been analyzed on the basis of the “fuzzy oil drop” model. The authors have determined the contribution of individual exon fragments to the creation of a common hydrophobic core and assessed the involvement of each chain in this process, depending on the number of complexed chains. Quantitative assessment of exons, chains, dimers, and the whole complex suggest that each of these units plays a different role in shaping the protein’s hydrophobic core.

Exceptionally high cumulative percentage of NUMTs originating from linear mitochondrial DNA molecules in the Hydra magnipapillata genome

BackgroundIn contrast to most animal genomes, mitochondrial genomes in species belonging to the phylum Cnidaria show distinct variations in genome structure, including the mtDNA structure (linear or circular) and the presence or absence of introns in protein-coding genes. Therefore, the analysis of nuclear insertions of mitochondrial sequences (NUMTs) in cnidarians allows us to compare the NUMT content in animals with different mitochondrial genome structures.ResultsNUMT identification in the Hydra magnipapillata, Nematostella vectensis and Acropora digitifera genomes showed that the NUMT density in the H. magnipapillata genome clearly exceeds that in other two cnidarians with circular mitochondrial genomes. We found that H. magnipapillata is an exceptional ancestral metazoan with a high NUMT cumulative percentage but a large genome, and its mitochondrial genome linearisation might be responsible for the NUMT enrichment. We also detected the co-transposition of exonic and intronic fragments within NUMTs in N. vectensis and provided direct evidence that mitochondrial sequences can be transposed into the nuclear genome through DNA-mediated fragment transfer. In addition, NUMT expression analyses showed that NUMTs are co-expressed with adjacent protein-coding genes, suggesting the relevance of their biological function.ConclusionsTaken together, our results provide valuable information for understanding the impact of mitochondrial genome structure on the interaction of mitochondrial molecules and nuclear genomes.

Structural role of exons in hemoglobin

Abstract The role of exons can be studied on many levels, one of which pertains to protein structure. It is a well-known fact that secondary structural motifs do not directly correspond to exons: helices, β-sheets and loops have all been identified as encoded by more than one exon. The relation between exon fragments and their involvement in shaping the three-dimensional (3D) structure of a protein body is subject to ongoing studies. In particular, the role of exons in stabilizing tertiary structures can be related to the structure of the hydrophobic core of the protein. Participation of specific polypeptide fragments (single exons) in hydrophobic stabilization reveals the role played by each fragment. In the course of the presented research, exons in selected proteins have been identified on the basis of GenBank files, imported from the nucleotide database at the National Center of Biotechnology Information. Amino acid sequences representing each exon were subsequently traced to parts of 3D structural forms. The participation of each exon fragment in shaping the hydrophobic core of the protein was measured using divergence entropy calculations. It was found that each protein contains at least one exon which encodes a structural fragment in accordance with the theoretical hydrophobic core model. This implies that the likely role of at least one exon in each protein is to generate a hydrophobic core which is, in turn, responsible for tertiary structural stabilization.

Polymorphisms on Chromosome 9p21 Confer a Risk for Acute Coronary Syndrome in a Chinese Han Population

BackgroundGenome-wide association studies have identified 2 single-nucleotide polymorphisms (SNPs) on chromosome arm 9p21, rs10757278, and rs2383207 that confer susceptibility to myocardial infarction. However, these data are mostly from Italian, American Caucasian, South Korean, and Japanese cohorts. This study is the first to investigate whether 6 SNPs (rs10757277, rs10757278, rs10757279, rs1333049, rs1333047, and rs10811656) are associated with acute coronary syndrome (ACS) in a Chinese Han population. MethodsWe performed a case-control analysis in 359 patients with ACS diagnosed by coronary angiography and 398 non-ACS controls of Han background between April 2007 and January 2008 to determine whether these 6 SNPs were associated with ACS. Exon fragments were genotyped by polymerase chain reaction–restriction fragment length polymorphism. ResultsAfter we adjusted for clinical parameters, we found the rs10757278 GG genotype to be associated with a significantly elevated risk of ACS (odds ratio [OR], 1.91; 95% confidence interval [CI], 1.35-2.68; P = 0.00035), the rs10811656 T allele to be associated with a higher risk of ACS (OR, 1.67; 95% CI, 1.26-2.23; P = 0.0016) than the CC genotype, and the rs1333047 TT genotype also to be associated with a higher risk of ACS (OR, 1.57; 95% CI, 1.15-2.06; P = 0.0052) than the CC and CT genotypes. After 14.2 ± 4.5 months of follow-up, the end-point data were obtained: death (cardiac and noncardiac), nonfatal myocardial infarction, and recurrent angina leading to repeated coronary angiography. We found that the rs10757278 GG genotype was significantly associated with recurrent angina compared with the AA and AG genotypes (P = 0.013). ConclusionsPolymorphisms on 9p21 were associated with ACS in a Chinese Han population. The rs10757278 GG genotype was further associated with adverse cardiac outcomes after ACS.

Disorder of sexual development in a Yorkshire terrier (78, XY; SRY-positive)

A 9-month-old Yorkshire terrier was admitted to the clinic because of abnormal sexual behaviour and clitoral hypertrophy. External examination confirmed standard development of caudal genital organs: vagina, vulva and cervix uteri. Serum profile of gonadotropin hormones 17 β-estradiol (<10.0 pg.ml(-1)) and testosterone (9.1 ng.ml(-1)) revealed the presence of testicular tissue. A midline laparotomy was performed to detect the cranial parts of the genital system. Gonads resembling testicles, structures indicating epididymis and rudimentary deferent ducts were resected, along with adherent part of the uterus. Cytogenetic analysis showed a male chromosomal complement 78, XY in all metaphases of the studied Yorkshire terrier dog. The chromosomal constitution was confirmed by fluorescence in situ hybridisation (FISH) with whole-chromosome painting probes specific for chromosomes X and Y, as well as by polymerase chain reaction (PCR) amplification of the 271-bp Y-linked fragment of SRY (the sex-determining region on the Y chromosome) gene. Sequencing of the dog's SRY gene coding region did not reveal any mutation. To search for potential mutation in the SOX9 gene (Sry-box containing gene 9), which is considered to be one of the key genes involved in the sex determination process, the PCR fragments of exons 1, 2 and 3 originating from the canine patient were sequenced in order to compare with both male and female healthy control dogs. In the analysed regions of the SOX9 gene, no mutation was found.

Variation in growth hormone (GH) of gene in exon sequence in three salmon types

We aimed to identify polymorphisms in the coding regions of the Salmo trutta caspius (s. t. caspius) growth hormone gene for comparison of the rate homology between sequences cited regards salmonids, because the variation between sequences of exons is very important at finally can change in the rate of expression of gene growth hormone, however this gene identified by marker genetics in salmonids, and in those of Salmo salar (s. salar) and salmo trutta (s. trutta). This single copy nuclear gene contains six exons, that the length of the exon fragments almost 1900 base pairs between nucleotide 35 and 1907. In s. t. caspius. Selective PCR reaction, and sequenced the products, that including fragments of from first to end of the GH gene. We also describe a novel polymorphism of the six exons fragment, these fragments indicated for studies regards evolution of fish GH genes, phylogeny of fishes, and genetic selection. The segments of GH exon were analyzed by DNAMAN program genetics. The results are shown, there were almost 90%, homology between the first three exons of S. t. caspius and S. salar accession number (AY614002.1) Regards other Salmons; S. salar (Accession number, M21573.1) and Rainbow trout there weren’t any homology, exon fragments second, third, fourth, fifth and sixth, the homology were high (almost 90-95 %), however between fragments of salmons, the rate of homology were high but the length of fragments between salmons were different. The length of fragments in s. salar more than other salmons including R. trout and S. t. caspius.

Comparison of 454 pyrosequencing methods for characterizing the major histocompatibility complex of nonmodel species and the advantages of ultra deep coverage

Characterization and population genetic analysis of multilocus genes, such as those found in the major histocompatibility complex (MHC) is challenging in nonmodel vertebrates. The traditional method of extensive cloning and Sanger sequencing is costly and time-intensive and indirect methods of assessment often underestimate total variation. Here, we explored the suitability of 454 pyrosequencing for characterizing multilocus genes for use in population genetic studies. We compared two sample tagging protocols and two bioinformatic procedures for 454 sequencing through characterization of a 185-bp fragment of MHC DRB exon 2 in wolverines (Gulo gulo) and further compared the results with those from cloning and Sanger sequencing. We found 10 putative DRB alleles in the 88 individuals screened with between two and four alleles per individual, suggesting amplification of a duplicated DRB gene. In addition to the putative alleles, all individuals possessed an easily identifiable pseudogene. In our system, sequence variants with a frequency below 6% in an individual sample were usually artefacts. However, we found that sample preparation and data processing procedures can greatly affect variant frequencies in addition to the complexity of the multilocus system. Therefore, we recommend determining a per-amplicon-variant frequency threshold for each unique system. The extremely deep coverage obtained in our study (approximately 5000×) coupled with the semi-quantitative nature of pyrosequencing enabled us to assign all putative alleles to the two DRB loci, which is generally not possible using traditional methods. Our method of obtaining locus-specific MHC genotypes will enhance population genetic analyses and studies on disease susceptibility in nonmodel wildlife species.

A rare heterozygous TRAF6 variant is associated with hypohidrotic ectodermal dysplasia

Mutations in the genes encoding components of the tumour necrosis factor (TNF)-α-like pathway cause hypohidrotic ectodermal dysplasia (HED). It has been postulated that the TNF receptor-associated factor 6 (TRAF6) is also involved in this pathway. To investigate mutations in the TRAF6 gene in an individual with HED. Genetic analysis was performed on TRAF6 in a patient with HED, her parents, her sister and 150 ethnically matched, healthy individuals. In the patient, sequencing analysis of one DNA strand revealed a deletion of eight nucleotides (c.1074-1081delCAATTTG) in the 5' fragment of the last exon of TRAF6, while no deletion was detected in the other DNA strand indicating a heterozygous mutation. No such sequence abnormality was detected in the patient's parents and her sister. This is the first report of a heterozygous TRAF6 sequence variant associated with symptoms typical of HED.

Elevated proline content in maize plants expressing a fragment of the proline dehydrogenase gene in antisense orientation

A fragment of the first exon of the proline dehydrogenase gene of arabidopsis (Arabidopsis thaliana L.) in antisense orientation was transferred to maize (Zea mays L.) genome by agrobacterial transformation in planta to elevate the content of free proline in maize cells. The presence of genetic structure carrying an antisense sequence of the fragment of the proline dehydrogenase gene (ASPG) from the genome of diploid maize seedlings was corroborated by PCR method. T-DNA insertions comprising ASPG were found in the genomes of 30 plants (1.2% of 2409 examined seedlings) belonging to the generation T0. A reliable 4.6-fold increase in proline content was registered in the leaves of transformed maize plants.

Comparison of immunohistochemistry, DNA sequencing and allele-specific PCR for the detection of IDH1 mutations in gliomas

Previous studies have identified mutations of the isocitrate dehydrogenase 1 (IDH1) gene in more than 70% of World Health Organization (WHO) grade II and III gliomas. The most frequent mutation leads to a specific amino acid change from arginine to histidine at codon 132 (c.395G>A, p.R132H). IDH1 mutated tumors have a better prognosis than IDH1 non-mutated tumors. The aim of our study was to evaluate and compare the methods of mIDH1 R132H immunohistochemistry, allele-specific PCR and DNA sequencing for determination of IDH1 status. We performed a retrospective study of 91 patients with WHO grade II (n=43) and III (n=48) oligodendrogliomas. A fragment of exon 4 spanning the sequence encoding the catalytic domain of IDH1, including codon 132, was amplified and sequenced using standard conditions. Allele-specific amplification was performed using two forward primers with variations in their 3' nucleotides such that each was specific for the wild-type or the mutated variant, and one reverse primer. Immunohistochemistry was performed with mouse monoclonal mIDH1 R132H. DNA was extracted from FFPE sections following macrodissection. IDH1 mutations were found in 55/90 patients (61.1%) by direct sequencing. R132H mutations were found in 47/55 patients (85.4%). The results of the allele-specific PCR positively correlated with those from DNA sequencing. Other mutations (p.R132C, p.R132S and pR132G) were found by DNA sequencing in 3, 3 and 2 tumors, respectively (8/55 patients, 14.6%). mIDH1 R132H immunostaining was found in the 47 patients presenting the R132H mutation (sensitivity 47/47, 100% for this mutation). None of the tumors presenting a wild-type IDH1 gene were stained (specificity 35/35, 100%). Our results demonstrate that immunohistochemistry using the mIDH1 R132H antibody and allele-specific amplification are highly sensitive techniques to detect the most frequent mutation of the IDH1 gene.

Genetic polymorphism in β-lactoglobulin gene of some sheep breeds in the Kingdom of Saudi Arabia (KSA) and its influence on milk composition

β-Lactoglobulin (β-Lg) is one of the most important proteins in mammals’ milk. It plays a crucial role in milk quality. The polymorphism of β-Lg gene can be used as a marker system. To analyze the genotype distribution of β-Lg gene in some sheep breeds reared in Taif region of Saudi Arabia and its influence on milk composition, sixty (60)animals belonging to four sheep breeds named Noami, Sawakni, Harry and Nagdi, were utilized. The polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) test was performed and genetic polymorphism was detected by the digestion of a 452 bp PCR fragment of exon II of β-Lg gene with the endonuclease RsaI. The results revealed that Noami and Sawakni breeds belong toβ-Lg-A genotype while Harry and Nagdi belong to genotype β-Lg-B. Sequence analysis of a 340 bp fragment in the promoter region of β-Lg showed polymorphismamong the examined breeds. Analysis of milk composition in the different breeds indicated that the total protein content of milk was the highest in Noami breed followed by Sawakni, Nagdi and Harry. Concerning milk total fat and total solids contents, Harry breed was the highest, while no significant difference was evident among different breeds in lactose or non-fat solid contents. These results indicate the feasibility of PCR-RFLP test for differentiating sheep breeds and the existence of a significant relationship between β-Lg-A genotype and total milk protein content while no clear association between β-Lg genotypes and other milk content was proved. Key words: Sheep, β-lactoglobulin, genetic polymorphism, PCR-RFLP, milk composition.

Polymorphism investigation of calpastatin gene in Zel sheep population of Iran by PCR-RFLP method

Meat tenderness is an importantquality characteristicfor which consumersare interested. Calpastatin is the calpain inhibitor enzyme and plays an important role in muscle growth and meat quality. The calpastin gene is located on sheep chromosome 5 and its polymorphisms are associated with economic traits. This study was performed to identify calpastatin gene polymorphisms using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. Random blood samples were collected from 100 Zel sheep. DNA was extracted by the salting out method and was amplified using PCR. The quantity and quality of the extracted DNA were determined by spectrophotometry and agarose gel electrophoresis. For amplification, a 622 bp fragment exon 1 of the calpastatin gene was used. Amplified fragments were digested by Msp1 restriction enzyme to determine the calpastatin genotypes, and two alleles. M and N were identified. The genotype frequencies of the MM, MN and NN were 0.62, 0.26 and 0.12, respectively in the population under study. Also, the allelic frequency of M and N were 0.75 and 0.25, respectively. Investigation of population genetic structure of Zel sheep revealed that it was not in Hardy-Weinberg equilibrium (p<0.05). The results indicate that it could be useful to consider genetic diversity at calpastatin locus in Zel sheep. Key words: Calpastatin gene, polymorphism, Zel sheep, meat and polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP).

Analysis of polymorphism within POU1F1 gene in relation to milk production traits in dairy Sarda sheep breed

The ovine POU1F1 gene is localized on chromosome 1 and it contains five introns and six exons. In different mammalian species some mutations in different exons are associated with different production traits. The aim of our research was to study the POU1F1 gene nucleotide sequence to detect possible polymorphisms and their relationships with milk productive traits in Sarda breed sheep. The study had been conducted on 140 ewes, 4 or 5 years old coming from a farm located in Sardinia. All the animals were multiparous, lactating and in their third to fifth lactation. Individual milk yield had been recorded monthly and for each sample fat, protein, casein, lactose, and somatic cell count values were analysed. A jugular blood sample was collected from each ewe to perform genomic DNA extraction. PCR, SSCP and sequencing analysis were carried out to examine the six exons to highlight possible SNPs. One-way ANOVA was used to analyse association of variants with milk yield and/or its composition. Two novel SNP were found: 121 C>T in the 5'UTR of the fourth intron fragment and 249 G>A in the 3'UTR of the sixth exon fragment. The statistical analysis did not shown association between milk productive traits and the found polymorphisms. However, further investigations about the promoter region or the prophet genes, like the PROP-1, could clarify its exact role in regulating the productive traits in sheep.

Genomic Analyses of Toll-like Receptor 4 and 7 Exons of Bos indicus from Temperate Sub-himalayan Region of India

Toll-like receptors (TLRs) play an important role in the recognition of invading pathogens and the modulation of innate immune responses in mammals. The TLR4 and TLR7 are well known to recognize the bacterial lipopolysaccharide (LPS) and single stranded (ssRNA) ligands, respectively and play important role in host defense against Gram-negative bacteria and ssRNA viruses. In the present study, coding exon fragments of these two TLRs were identified, cloned, sequenced and analyzed in terms of insertion-deletion polymorphism, within bovine TLRs 4 and 7, thereby facilitating future TLR signaling and association studies relevant to bovine innate immunity. Comparative sequence analysis of TLR 4 exons revealed that this gene is more variable, particularly the coding frame (E3P1), while other parts showed percent identity of 95.7% to 100% at nucleotide and amino acid level, respectivley with other Bos indicus and Bos taurus breeds from different parts of the world. In comparison to TLR4, sequence analysis of TLR7 showed more conservation among different B. indicus and B. taurus breeds, except single point mutation at 324 nucleotide position (AAA to AAM) altering a single amino acid at 108 position (K to X). Percent identity of TLR7 sequences (all 3 exons) was between 99.2% to 100% at nucleotide and amino acid level, when compared with available sequence database of B. indicus and B. taurus. Simple Modular Architecture Research Tool (SMART) analysis showed variations in the exon fragments located in the Leucine Rich Repeat (LRR) region, which is responsible for binding with the microbial associated molecular patterns and further, downstream signaling to initiate anti-microbial response. Considering importance of TLR polymorphism in terms of innate immunity, further research is warranted.

Expression of Phosphophoryn Is Sufficient for the Induction of Matrix Mineralization by Mammalian Cells

Mineralized tissues such as dentin and bone assemble extracellular matrices uniquely rich in a variety of acidic phosphoproteins. Although these proteins are presumed to play a role in the process of biomineralization, key questions regarding the nature of their contributions remain unanswered. First, it is not known whether highly phosphorylated proteins alone can induce matrix mineralization, or whether this activity requires the involvement of other bone/dentin non-collagenous proteins. Second, it remains to be established whether the protein kinases that phosphorylate these acidic proteins are unique to cells responsible for producing mineralized tissues. To begin to address these questions, we consider the case of phosphophoryn (PP), due to its high content of phosphate, high affinity for Ca(2+), and its potential role in hydroxyapatite nucleation. We have created a model system of biomineralization in a cellular environment by expressing PP in NIH3T3 fibroblasts (which do not produce a mineralized matrix); as a positive control, PP was expressed in MC3T3-E1 osteoblastic cells, which normally mineralize their matrices. We show that expression of PP in NIH3T3 cells is sufficient for the induction of matrix mineralization. In addition, assessment of the phosphorylation status of PP in these cells reveals that the transfected NIH3T3 cells are able to phosphorylate PP. We suggest that the phosphorylation of PP is essential for mineral formation. The principle goal of this study is to enrich the current knowledge of mineralized tissue phosphorylation events by analyzing them in the context of a complete cellular environment.

Detection of IDH1 mutations in a series of 91 oligodendrogliomas: Comparison of immunohistochemistry, DNA sequencing, and allele-specific PCR.

2020 Background: IDH1 mutations have been indentified in about 70% of WHO grade 2 and 3 astrocytomas and oligodendrogliomas, conferring a better prognosis. The most frequent mutation leads to a specific amino-acid change from arginine to histidine at codon 132 (c.395G>A, p.R132H). We compared mIDH1 R132H immunohistochemistry, allele specific PCR and DNA sequencing for determination of IDH1 status. Methods: We performed a retrospective study of 91 WHO grade 2 (n=43; mean 40.6 years) and grade 3 (n=48; mean 50 years) oligodendrogliomas, diagnosed in the Neurosurgery Department of Nantes. Immunohistochemistry was performed with mouse monoclonal mIDH1 R132H (gift von Dr von Deimling, Heidelberg, Germany). DNA was extracted from FFPE sections following macrodissection using an iPrep system. A fragment of exon 4 (122 bp) spanning the sequence encoding the catalytic domain of IDH1 including codon 132 was amplified and sequenced using standard conditions. Allele specific amplification was performed using forward primers with variations in their 3′ nucleotides such that each was specific for the wild type or the mutated variant. Results: DNA could be amplified in 90 cases. IDH1 mutations were found in 55/90 patients (61.1%) by direct sequencing. Among these patients, R132H mutations was found in 47/55 patients (85.4%). The results of the allele specific PCR was totally correlated to DNA sequencing. Other mutations (p.R132C, p.R132S and pR132G) were found by DNA sequencing in 3, 3 and 2 tumors, respectively (8/55 patients, 14.6%). mIDH1 R132H immunostaining was found in the 47 patients presenting the R132H mutation (sensitivity 47/47 = 100% for this mutation). None of the tumors presenting a wild type IDH1 gene were stained (specificity 35/35 = 100%). Conclusions: Immunohistochemistry with mIDH1 R132H antibody is feasible, even in case of small biopsies, highly sensitive and specific. Because it only recognizes the most frequent mutation of IDH1 gene, immunohistochemical negative cases must then be tested by allele-specific PCR or DNA sequencing.

Activity-Dependent Transcription of BDNF Enhances Visual Acuity during Development

In the developing Xenopus tadpole, conditioning with 20 min of visual stimulation leads to increased proBDNF protein levels in the tectum measured 4 hr later. Following conditioning, the ability to induce direction selectivity in tectal neurons, as well as both retinotectal long-term potentiation and depression, thought to underlie this phenomenon, was strongly facilitated. This facilitation was blocked by knockdown of BDNF expression in tectal neurons. Animals that had been exposed to visual conditioning and subsequently received normal visual input for 7-11 hr exhibited higher spatial frequency thresholds of tectal cell responses to counterphasing gratings than nonconditioned control animals. An improvement in visual acuity was confirmed by enhanced sensitivity to counterphasing gratings in a behavioral test. These results indicate that brief sensory stimulation, by initiating nuclear transcription and de novo protein synthesis of BDNF, can facilitate the refinement of response properties in the developing visual system.

Mutations in exons 5, 7 and 8 of the human voltage-dependent anion channel type 3 (VDAC3) gene in sperm with low motility

Summary Voltage-dependent anion channels (VDACs1-3) are pore-forming proteins located in the outer mitochondrial membrane of various tissues and in human sperm flagella. VDACs are involved in metabolite fluxes between mitochondria and other cell compartments and play a role in the motility of mammalian spermatozoa. VDAC3-deficient male mice lacking exons 5–8 were reported to be healthy but infertile (Sampson et al., 2001). We analysed mutations in these exons of the human VDAC3 (hVDAC3) gene in spermatozoa of 32 asthenozoospermic patients. Previously, we reported on nucleotide substitution mutations in exon 6 (Asmarinah et al., 2005); now, we continue with the examination of exons 5, 7 and 8. Amplification of the exon fragments of the hVDAC3 gene was carried out by polymerase chain reaction (PCR) using specific primers for each exon followed by sequencing. We found various types of mutations in the examined exons of the hVDAC3 gene, such as deletion and nucleotide substitution in spermatozoa from seven of the 30 asthenozoospermic sperm samples (and unconfirmed insertions in two other ones) but none in spermatozoa from normozoospermic controls. Our preliminary data, although of small numbers investigated, suggest that genetic mutations in the hVDAC3 gene could be an explanation for the aetiology of infertility in these asthenozoospermic patients.

Characterization of mannose-binding lectin plasma levels and genetic polymorphisms in HIV-1-infected individuals

The present study investigated the association between mannose-binding lectin (MBL) gene polymorphism and serum levels with infection by HIV-1. Blood samples (5 mL) were collected from 97 HIV-1-infected individuals resident in Belém, State of Pará, Brazil, who attended the Special Outpatient Unit for Infections and Parasitic Diseases (URE-DIPE). CD4+ T-lymphocyte count and plasma viral load were quantified. A 349bp fragment of exon 1 of the MBL was amplified via PCR, using genomic DNA extracted from controls and HIV-1-infected individuals, following established protocols. MBL plasma levels of the patients were quantified using an enzyme immunoassay kit. Two alleles were observed: MBL*O, with a frequency of 26.3% in HIV-1-infected individuals; and the wild allele MBL*A (73.7%). Similar frequencies were observed in the control group (p > 0.05). Genotype frequencies were distributed according to the Hardy-Weinberg equilibrium in both groups. Mean MBL plasma levels varied by genotype, with statistically significant differences between the AA and AO (p < 0.0001), and AA and OO (p < 0.001) genotypes, but not AO and OO (p = 0.17). Additionally, CD4+ T-lymphocytes and plasma viral load levels did not differ significantly by genotype (p > 0.05). The results of this study do not support the hypothesis that MBL gene polymorphism or low plasma MBL concentrations might have a direct influence on HIV-1 infection, although a broader study involving a large number of patients is needed.

Analysis of the ORF region of the prion protein gene in the Botosani Karakul sheep breed from Romania

Ovine scrapie is considered a neurodegenerative disease characterized by the accumulation of an abnormal, protease-resistant isoform of PrP. The polymorphisms at codons 136, 154, and 171 are associated with susceptibility to scrapie. In order to detect the existing polymorphisms at the 3 codons we used the real-time PCR technique. The obtained genotypes were confirmed by PCR amplification and sequencing of a 402 bp fragment of exon 3 of the prion gene. Known polymorphisms were detected at codons 136, 154, and 171 and 5 different genotypes were determined. The results showed that all animals were homozygous for A at codon 136 and homozygous for R at codon 154 and 3 out of 4 previously identified polymorphisms at residue 171 were detected (R, H, and Q). Two new polymorphisms were detected at codon 189 and 199, as well as 2 other reported polymorphisms at codons 141 and 143. The frequency of high risk ARQ/ARQ genotype was about 47%. The aim of this study was to determine same preliminary information on the PrP gene-associated susceptibility in scrapie in the Botosani Karakul sheep breed.