scholarly journals Characterization of a New Alloantigen (SH) on the Human Neutrophil Fcγreceptor IIIb

Blood ◽  
1997 ◽  
Vol 89 (3) ◽  
pp. 1027-1034 ◽  
Author(s):  
Juergen Bux ◽  
Ernst-Ludwig Stein ◽  
Philippe Bierling ◽  
Patricia Fromont ◽  
Mary Clay ◽  
...  
Keyword(s):  
Nucleotide Sequence Analysis ◽  
Polymorphism Analysis ◽  
Mutational Event ◽  
Coding Region ◽  
Specific Pcr ◽  
Pcr Products ◽  
Polymerase Chain ◽  
Antigen Frequency ◽  
Neonatal Neutropenia

Abstract Polymorphic structures of the neutrophil Fcγreceptor IIIb (FcγRIIIb) result in alloantibody formation that causes alloimmune neonatal neutropenia and transfusion reactions. Alloantigens located on FcγRIIIb include the antigens NA1 and NA2. In four cases of alloimmune neonatal neutropenia, granulocyte-specific alloantibodies directed against a thus far unknown antigen were detected by granulocyte agglutination and immunofluorescence tests in the maternal sera. By the use of the monoclonal antibody–specific immobilization of granulocyte antigens (MAIGA) assay, the new antigen, termed SH, was located on the FcγRIIIb. Nucleotide sequence analysis of the FcγRIIIb coding region from a SH(+) individual showed a single-base C→A mutation at position 266, which results in an Ala78Asp amino acid substitution. A family study confirmed that this nucleotide difference is inherited, and corresponds to the SH phenotype. Serologic typing of 309 randomly selected individuals showed an antigen frequency of 5% in the white population. The same frequency was found by genotyping, for which a technique based on polymerase chain reaction (PCR) using sequence-specific primers (PCR-SSP) was developed. Typing of all SH(+) individuals for NA1 and NA2, and PCR-restriction fragment length polymorphism analysis of the NA-specific PCR products from five SH(+) individuals using the SH-specific endonuclease SfaN I showed that SH antigen is very probably the result of an additional mutational event in the NA2 form of the FcγRIIIB gene. Immunochemical studies also demonstrated that the SH determinants reside on the 65- to 80-kD NA2 isoform of the FcγRIIIb. Our findings show the existence of an additional polymorphism of the FcγRIIIb, which can result in alloantibody formation causing alloimmune neonatal neutropenia.

Characterization of the Original Christmas Disease Mutation (Cysteine 206 → Serine): From Clinical Recognition to Molecular Pathogenesis

1992 ◽  
Vol 67 (01) ◽  
pp. 063-065 ◽  
Author(s):  
Sherryl A M Taylor ◽  
Jacalyn Duffin ◽  
Cherie Cameron ◽  
Jerome Teitel ◽  
Bernadette Garvey ◽  
...  
Keyword(s):  
Nucleotide Substitution ◽  
Novel Mutation ◽  
Factor Ix ◽  
Causative Mutation ◽  
Coding Region ◽  
Body Of Knowledge ◽  
Polymerase Chain ◽  
Splice Junctions

SummaryChristmas disease was first reported as a distinct clinical entity in two manuscripts published in 1952 (1, 2). The eponym associated with this disorder, is the surname of the first patient examined in detail and reported by Biggs and colleagues in a paper describing the clinical and laboratory features of seven affected individuals (3). This patient has severe factor IX coagulant deficiency (less than 0.01 units/ml) and no detectable circulating factor IX antigen (less than 0.01 units/ml). Coding sequence and splice junctions of the factor IX gene from this patient have been amplified in vitro through the polymerase chain reaction (PCR). One nucleotide substitution was identified at nucleotide 30,070 where a guanine was replaced by a cytosine. This mutation alters the amino acid encoded at position 206 in the factor IX protein from cysteine to serine. The non conservative nature of this substitution, the absence of this change in more than 200 previously sequenced factor IX genes and the fact that the remainder of the coding region of this gene was normal, all provide strong circumstantial evidence in favour of this change being the causative mutation in this patient. The molecular characterization of this novel mutation in the index case of Christmas disease, contributes to the rapidly expanding body of knowledge pertaining to Christmas disease pathogenesis.

scholarly journals First Report of a Group 16SrII Phytoplasma Infecting Chickpea in Oman

Plant Disease ◽  
2006 ◽  
Vol 90 (7) ◽  
pp. 973-973 ◽  
Author(s):  
N. A. Al-Saady ◽  
A. M. Al-Subhi ◽  
A. Al-Nabhani ◽  
A. J. Khan
Keyword(s):  
Nested Pcr ◽  
Animal Feed ◽  
Restriction Enzymes ◽  
Universal Primers ◽  
Polymorphism Analysis ◽  
Disease Symptoms ◽  
First Report ◽  
Pcr Products ◽  
Polymerase Chain ◽  
Group 2

Chickpea (Cicer arietinum), locally known as “Dungo”, is grown for legume and animal feed mainly in the interior region of Oman. During February 2006, survey samples of chickpea leaves from plants showing yellows disease symptoms that included phyllody and little leaf were collected from the Nizwa Region (175 km south of Muscat). Total nucleic acid was extracted from asymptomatic and symptomatic chickpea leaves using a cetyltrimethylammoniumbromide method with modifications (3). All leaf samples from eight symptomatic plants consistently tested positive using a polymerase chain reaction assay (PCR) with phytoplasma universal primers (P1/P7) that amplify a 1.8-kb phytoplasma rDNA product and followed by nested PCR with R16F2n/R16R2 primers yielding a product of 1.2 kb (2). No PCR products were evident when DNA extracted from healthy plants was used as template. Restriction fragment length polymorphism analysis of nested PCR products by separate digestion with Tru9I, HaeIII, HpaII, AluI, TaqI, HhaI, and RsaI restriction enzymes revealed that a phytoplasma belonging to group 16SrII peanut witches'-broom group (2) was associated with chickpea phyllody and little leaf disease in Oman. Restriction profiles of chickpea phytoplasma were identical with those of alfalfa witches'-broom phytoplasma, a known subgroup 16SrII-B strain (3). To our knowledge, this is the first report of phytoplasma infecting chickpea crops in Oman. References: (1) A. J. Khan et al. Phytopathology, 92:1038, 2002. (2). I.-M. Lee et al. Int. J. Syst. Bacteriol. 48:1153, 1998 (3) M. A. Saghai-Maroof et al. Proc. Natl. Acad. Sci. USA. 81:8014, 1984.

scholarly journals Sex Determination During Inflorescence Bud Differentiation in Monoecious Pistacia chinensis Bunge

Forests ◽  
2019 ◽  
Vol 10 (3) ◽  
pp. 202 ◽  
Author(s):  
Qian Bai ◽  
Chenyi Zhu ◽  
Xia Lei ◽  
Tao Cao ◽  
Shuchai Su ◽  
...  
Keyword(s):  
Sex Determination ◽  
Early Stage ◽  
Sex Differentiation ◽  
Banding Patterns ◽  
Vegetative Period ◽  
Pcr Products ◽  
Polymerase Chain ◽  
Sex Determination Mechanisms

Pistacia chinensis Bunge is widely acknowledged to be dioecious, but rare monoecious individuals have been found. However, the origin of monoecism and the sex differentiation of different sex types remain intriguing questions. Here, sex expressions were explored by identification of sex-associated DNA markers, determination of the sex stability after grafting, and histological characterization of inflorescence bud development using anatomical analysis. The results showed that (1) although polymorphisms among individuals existed, the banding patterns of Polymerase Chain Reaction (PCR) products for different sex types on the same monoecious tree were consistent; (2) the sex expressions of grafted trees were not consistent with those of scions, indicating that monoecism probably did not originate from a stable bud mutation; and (3) both males and females underwent a bisexual period, then the stamen primordia in female buds degenerated into the second round tepals, while the pistil primordia in male buds gradually disappeared. During the sex differentiation phase, female buds were spindle-shaped, while the male buds were full teardrop-shaped, and male buds were bigger than female buds. Taken together, no sex-associated DNA marker was found, sex expressions were unstable after grafting, and the alternative sex organs appeared in the early stage of sex differentiation, suggesting that sex determination occurred during floral development instead of the early vegetative period. These results indicated that the sex expressions may be affected by environmental factors, increasing the understanding of sex determination mechanisms in P. chinensis and other species.

scholarly journals Morphological and molecular characterization of Strongyloides ophidiae (Nematoda, Strongyloididae)

2009 ◽  
Vol 84 (2) ◽  
pp. 136-142 ◽  
Author(s):  
K.R. dos Santos ◽  
B.C. Carlos ◽  
K.S. Paduan ◽  
S.M. Kadri ◽  
T.H. Barrella ◽  
...  
Keyword(s):  
Dna Analysis ◽  
Dna Amplification ◽  
Strongyloides Stercoralis ◽  
Morphological Data ◽  
Nucleotide Sequence Analysis ◽  
Parasitic Female ◽  
Strongyloides Venezuelensis ◽  
Polymerase Chain ◽  
Morphological And Molecular Characterization

AbstractThe aim of the present study is to report morphological data from parasitic female, rhabditoid and filarioid larvae, free-living female worms and eggs of Strongyloides ophidiae (Nematoda, Strongyloididae). In addition, a molecular DNA analysis was carried out using a pool of eight S. ophidiae parasitic females. Samples were obtained from the small intestine of Oxyrhopus guibei (Serpentes, Colubridae) collected in the municipality of Lençóis Paulista, State of São Paulo, Brazil. DNA amplification by polymerase chain reaction (PCR) resulted in a 350 bp band for samples containing S. ophidiae and Strongyloides venezuelensis DNA. Strongyloides ophidiae nucleotide sequence analysis showed 98% similarity with Strongyloides procyonis and 97% with Strongyloides cebus, Strongyloides stercoralis, Strongyloides fuelleborni and Strongyloides sp. from snakes.

Estimating the prevalence of mixed-type gonococcal infections in Queensland, Australia

Sexual Health ◽  
2015 ◽  
Vol 12 (5) ◽  
pp. 439 ◽  
Author(s):  
Ella Trembizki ◽  
Christine Doyle ◽  
Cameron Buckley ◽  
Amy Jennison ◽  
Helen Smith ◽  
...  
Keyword(s):  
Sequence Data ◽  
Nucleic Acid Amplification ◽  
Clinical Samples ◽  
Anatomical Site ◽  
Diagnostic Screening ◽  
Specific Pcr ◽  
Screening Population ◽  
Pcr Products ◽  
Polymerase Chain ◽  
The One

Background Mixed gonococcal infections within the one anatomical site have been recognised but questions remain over how often they occur. In this study, the aim was to estimate the prevalence of mixed gonococcal infections using novel real-time polymerase chain reaction (PCR) methods that were developed and validated, targeting the gonococcal porB gene. Methods: Neisseria gonorrhoeae strains were categorised into three different porB groups, based on sequence data derived from N. gonorrhoeae multi-antigen sequence typing (NG-MAST) analyses of local isolates. Specific PCR methods for each group were then developed and these PCR methods were used to test clinical samples (n = 350) that were positive for gonorrhoea as determined by nucleic acid amplification test (NAAT) diagnostic screening. Results: Initial validation using isolates showed the group PCR methods proved 100% sensitive and 100% specific for their respective porB groups. When applied to the clinical specimens, 298/350 (85%) provided positive results by the group PCR methods. Of these, four specimens showed evidence of mixed infections, supported by subsequent DNA sequencing of the PCR products. Conclusions: The data provide further evidence of mixed gonococcal infections at the same anatomical site, but show that such infections may be relatively infrequent (1.3%; 95% confidence interval 0.01–2.6%) in a general screening population.

scholarly journals Genetic and Morphological Characterization of Colletotrichum acutatum Causing Anthracnose of Lupins

2002 ◽  
Vol 92 (9) ◽  
pp. 986-996 ◽  
Author(s):  
Pedro Talhinhas ◽  
S. Sreenivasaprasad ◽  
João Neves-Martins ◽  
Helena Oliveira
Keyword(s):  
Morphological Characters ◽  
Morphological Characterization ◽  
Colletotrichum Acutatum ◽  
Distinct Subgroup ◽  
Specific Pcr ◽  
Pathogen Diversity ◽  
Polymerase Chain ◽  
Arbitrarily Primed Pcr ◽  
Pathogen Diagnosis

Anthracnose, caused by Colletotrichum sp., is a serious problem of lupins (Lupinus spp.) worldwide. Morphological characters and molecular markers were used to characterize 43 Colletotrichum isolates from lupins, 8 isolates from other hosts, and 18 reference isolates representing related Colletotrichum spp., to assess the pathogen diversity and resolve its taxonomy. All lupin Colletotrichum isolates tested positive with C. acutatum-specific polymerase chain reaction (PCR) and did not test positive with C. gloeosporioides-specific PCR. Spore shape and colony diameter as well as insensitivity to benomyl grouped the lupin anthracnose isolates closer to C. acutatum than to C. gloeosporioides. Analysis of internal transcribed spacer (ITS) sequences of 57 Colletotrichum isolates grouped all lupin isolates with C. acutatum and distinct from C. gloeosporioides. Further, tub2 and his4 sequences revealed groups concordant with ITS, reducing the excessive dependence on the latter. Arbitrarily primed-PCR and amplified fragment length polymorphism analyses revealed intraspecific subgroups, but neither was useful to decipher species level relationships. ITS, tub2, and his4 results strongly support designating lupin anthracnose pathogen as C. acutatum or its subspecies. Most Colletotrichum isolates from lupins from worldwide locations are genetically homogeneous and form a distinct subgroup within C. acutatum. Present results also underline the potential of the C. acutatum-specific PCR for routine pathogen diagnosis.

scholarly journals Molecular Characterization of a Re-Emergent Brugia Malayi Parasite in Sri Lanka, Suggestive of a Novel Strain With Close Nucleotide Homology to Brugia Pahangi

2020 ◽  
Author(s):  
Chandana Harendra Mallawarachchi ◽  
Nilmini T. G. A. N Chandrasena ◽  
Ranjan Premarathna ◽  
S.M.N.S.M. Maleesha Mallawarachchi ◽  
Nilmini Y. I. S. Gunawardane ◽  
...  
Keyword(s):  
Sri Lanka ◽  
Its2 Region ◽  
Species Identity ◽  
Internal Transcribed Spacer Region ◽  
Region Gene ◽  
Specific Pcr ◽  
Baited Traps ◽  
Human Blood Samples ◽  
Polymerase Chain

Abstract Background Brugian filariasis has re-emerged in Sri Lanka after four decades of quiescence. As microscopy alone was insufficient for ascertaining the species identity of the re-emerged sub-periodic Brugia spp. parasite, molecular speciation was performed. The transmission dynamics of the parasite was studied by entomological procedures.Methods Human blood samples positive for Brugia spp. microfilariae (MF) (n=8) were collected and DNA extracted using ReliaPrep™ Blood DNA Miniprep System (modified). Polymerase chain reaction (PCR) was performed with pan-filarial primers specific for the internal transcribed spacer region 2 (ITS2) of the ribosomal DNA (rDNA) of MF. Results Of those tested, seven (87.5%) yielded a band at 615bp establishing the species identity of the re-emerged filarial parasite as B. malayi. Comparison of the ITS2 region gene sequences of B. malayi MF isolated from humans (n=2), dogs (n=3) and cats (n=6) with GenBank sequences revealed a higher sequence homology with B. pahangi than B. malayi, but phylogeny was closer to B. malayi. A total of 82 mosquitoes of genus Mansonia comprising of M. annulifera (65), M. uniformis (14) and M. indiana (3) were collected by cattle-baited traps. Mosquito dissections identified 17 infected mosquitoes: one M. uniformis (7.14%) and 16 M. annulifera (24.6%). The DNA extracts of all infected Mansonia mosquitoes elicited the 615bp band on pan-filarial primer specific PCR. Conclusions The re-emergent B. malayi is a genetic variant or a novel species closely related to B. malayi and B. pahangi. Mansonia spp. mosquitoes were vectors of this zoonotic variant B. malayi circulating among cats, dogs and humans in Sri Lanka.

scholarly journals Characterization of clone-specific rearrangement T-cell receptor gamma- chain genes in lymphomas and leukemias by the polymerase chain reaction and DNA sequencing

Blood ◽  
1994 ◽  
Vol 84 (2) ◽  
pp. 574-581 ◽  
Author(s):  
M Kneba ◽  
I Bolz ◽  
B Linke ◽  
J Bertram ◽  
D Rothaupt ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Polymerase Chain Reaction ◽  
T Cell ◽  
Dna Sequencing ◽  
Cell Lines ◽  
Chain Reaction ◽  
Gamma Chain ◽  
Pcr Products ◽  
Polymerase Chain

The structures of rearranged gamma-chain T-cell antigen receptor (TCR) genes were analyzed in 5 cases of T-cell acute lymphoblastic leukemia (T-ALL), in 15 cases of peripheral T-cell non-Hodgkin's lymphoma (T- NHL), in 1 case with large granular CD8 lymphocytosis, 1 case with CD8 lymphocytosis after autologous bone marrow transplantation for Hodgkin's disease, and in 2 cases with nonneoplastic diseases. Rearranged V-J TCR gamma-gene segments were amplified by the polymerase chain reaction (PCR). Because most of the biopsy tissue or bone marrow samples contained significant amounts of admixed nonmalignant T-cells, direct DNA sequencing of the PCR products yielded mixed sequence data because of coamplification of clonal together with polyclonal TCR gamma V-N-J junctions. Reliable data could only be obtained by cloning the V gamma-J gamma PCR products and sequencing several (4 to 10) randomly chosen clones. In the polyclonal samples, all PCR-derived clones differed in their specific V-N-J junctions, as expected. In the two T- cell lines and in most of the T-cell malignancies, monoclonal PCR products could be identified by the demonstration of clonally restricted V-N-J junctions. In most cases, this information yielded the desired clone-specific sequence and showed a background population of polyclonal TCR gamma cells in each specimen, except for those that were obtained from the T-ALL samples, the cell lines, or the NHL samples with high tumor cell fraction. The results obtained by PCR-directed sequencing were confirmed by temperature-gradient gel electrophoresis (TGGE) that showed distinct DNA bands only with the PCR products containing predominant (ie, monoclonal) TCR gamma V-N-J junctions. By combined sequence and TGGE analysis, it was found that PCR/TGGE is able to distinguish between monoclonal and polyclonal TCR gamma-PCR products. This finding prompted us to complete the analysis of the TCR gamma locus in the samples by PCR/TGGE using primer mixes which covered all possible V gamma and J gamma recombinations. Monoclonality was shown with all mixes by PCR/TGGE in 21 of 24 (87%) of the lymphoproliferations. In summary, the present study shows that the combination of amplifying TCR gamma V-N-J junctions by PCR with the identification of clonal PCR products by TGGE and DNA sequencing is a reliable method for the characterization of clonal TCR gamma sequences.

scholarly journals Molecular analysis of glycophorin C deficiency in human erythrocytes

Blood ◽  
1993 ◽  
Vol 81 (10) ◽  
pp. 2799-2803
Author(s):  
R Winardi ◽  
M Reid ◽  
J Conboy ◽  
N Mohandas
Keyword(s):  
Mechanical Stability ◽  
Mutant Gene ◽  
Erythrocyte Shape ◽  
Normal Individuals ◽  
Pcr Products ◽  
Protein Isoform ◽  
Polymerase Chain ◽  
Exon 4 ◽  
Glycophorin C

Human erythrocyte glycophorin C plays a functionally important role in maintaining erythrocyte shape and regulating membrane mechanical stability. We report here the characterization of the glycophorins C and D deficiency in erythrocytes of the Leach phenotype. Glycophorin C gene is encoded by 4 exons. Amplification of reticulocyte cDNA from Leach phenotype and normal individuals generated a 140-bp fragment when using primers spanning exons 1 and 2. However, no polymerase chain reaction (PCR) products were detected in the Leach phenotype using primers flanking either exons 1 and 3 or exons 1 and 4, suggesting that the 3' end of the mRNA was missing or altered. Exon 4 also appeared to be missing from Leach genomic DNA, based on both Southern hybridization and PCR. These results indicate that an absence of glycophorin C and glycophorin D in erythrocytes from these Leach phenotype individuals is a consequence of a deletion or marked alteration of exon 3 and exon 4 of their glycophorin C gene. Surprisingly, the mutant gene encodes an mRNA stable enough to be detected in circulating reticulocytes. Although this mRNA could encode an N-terminal fragment of glycophorin C, these protein isoform(s) would not be expressed in the membrane because they lack the transmembrane and cytoplasmic domains.

Sign in / Sign up

Export Citation Format

Share Document