Objective: To analyze the effect of the identification and evaluation of Escherichia (E.) coli and Shigella, based on the upstream flanking sequences of CRISPR1. Methods: Both CRISPR and cas sequences were obtained through the BLAST with repeating sequences against the publicly complete genome in GenBank that related to E. coli and Shigella. Clustal X was used to perform multi-sequences alignment of the flanking sequences. PCR method was used to amplify the upstream flanking sequences of CRISPR1 in order to appraise the effect of identification and evaluation of upstream flanking sequences on E. coli and Shigella, which were based on the upstream flanking sequences of CRISPR1. Results: The results showed that 73.4% of the strains containing the I-E CRISPR/Cas that belonged to the phylogroups A, B1, D while 8.4% strains carried the I-F CRISPR/Cas. Another 17.2% of the strains owned CRISPR3-4 (non-CRISPR/Cas) only belonged to the phylogroups B2. All the Shigella strains carried I-E CRISPR/Cas. More than 99% of similarity the CRISPR1 upstream-flanking sequences was seen in E. coli (except B2) and Shigella and E. coli (B2). Both sensitivity and specificity were greater than 91% after PCR amplification in the region to identify the E.coli and Shigella. Conclusion: The upstream of CRISPR1 could achieve a preliminary identification effect on E.coli and Shigella.
Clustered Regularly Interspaced Short Palindromic Repeats/genetics* ; DNA, Bacterial/genetics* ; Escherichia coli/isolation & purification* ; Genotype ; Humans ; Molecular Sequence Data ; Sequence Analysis, DNA ; Shigella/isolation & purification*

OBJECTIVETo explore the serological characteristics and molecular basis for an individual with para-Bombay phenotype.

METHODSBlood type of the proband was determined with routine serological methods. Exons 6 and 7 of the ABO gene and coding regions of the FUT1 and FUT2 genes were amplified by PCR and sequenced.

RESULTSThe para-Bombay phenotype was confirmed to be of Ah-secretion type. The genotype of the individual was determined as A102/O01. Position 328 of the FUT1 gene was mutated from A to G, resulting in replacement of Alanine (Ala) at position 110 by Threonine (Thr).

CONCLUSIONThe G to A mutation of nt328 of the FUT1 gene probably underlies the para-Bombay phenotype in this individual.

ABO Blood-Group System ; genetics ; Adult ; Alleles ; Base Sequence ; Exons ; Female ; Genotype ; Humans ; Molecular Sequence Data ; Mutation ; Point Mutation

OBJECTIVETo analyze a sample with ABO subgroup using serological and molecular methods.

METHODSThe ABO phenotype of the sample was determined with a tube method, and the activity of glycosyltransferases was determined with an uridine diphosphate galactose transferring method. The ABO gene of the propositus was identified by PCR with sequence-specific primers (PCR-SSP). In addition, exons 6 and 7 of the ABO gene were cloned and sequenced.

RESULTSNeither A nor B antigen was identified in the propositus, despite that its anti-B antibody was found to be attenuated. No activity of α -1, 3-D-galactosyltransferase was detected in the serum. The presence of B and O alleles were confirmed by PCR-SSP, and a novel mutation (562C to T) of the exon 7 was confirmed by sequencing, which has led to an amino acid substitution (Arg to Cys) at position 188. The genotype of the propositus was determined as Bnew/O.

CONCLUSIONA novel B allele has been identified, which was named as Bw39 by the Blood Group Antigen Gene Mutation Database (BGMUT).

ABO Blood-Group System ; genetics ; Adult ; Alleles ; Amino Acid Substitution ; Base Sequence ; Exons ; Galactosyltransferases ; genetics ; Humans ; Male ; Molecular Sequence Data ; Point Mutation

OBJECTIVETo explore the molecular basis of an individual with Bel variant of the ABO blood group.

METHODSThe ABO antigen and serum antibody of the individual were detected by serological method. All coding regions and flanking introns of the ABO gene were amplified with PCR and sequenced bidirectionally. The haplotypes of the individual were analyzed by cloning and sequencing. A three dimensional model of the mutant protein was constructed and analyzed.

RESULTSThe individual has expressed a very weak B antigen on its red blood cells by absorption and elution testing, which was identified as a Bel variant phenotype. The heterozygous sites in exon 6 (261del/G) and exon 7 (297A/G, 484del/G, 526C/G, 657C/T, 703G/A, 796C/A, 803G/C, 930G/A) of the coding region of the ABO gene were identified by direct sequencing. Haplotype analysis showed that the individual has carried an O01 allele and a novel B allele. The sequence of the novel B allele was identical to B101 except for a del G at nucleotide position 484 (484delG), which was nominated as B120 by the Blood Group Antigen Gene Mutation Database (dbRBC NCBI). The 484delG mutation of the B allele has led to a reading frame shift and created a premature terminal codon for the glycosyltransferase (GT) enzyme. Prediction of the 3D structure suggested that the GT enzyme has become an incomplete protein only with its N-terminal region.

CONCLUSIONThe 484delG mutation of the glycosyltransferase B gene has probably abolished or reduced the enzymatic activity and resulted in the Bel variant phenotype.

ABO Blood-Group System ; genetics ; Alleles ; Base Sequence ; Exons ; Female ; Genotype ; Glycosyltransferases ; genetics ; Humans ; Molecular Sequence Data ; Mutation ; Sequence Deletion

OBJECTIVETo detect potential mutation of EXT1 gene in a pedigree affected with multiple osteochondroma and explore its pathogenic mechanism.

METHODSThe coding regions and their flanking sequences of the EXT1/EXT2 genes were subjected to PCR amplification and Sanger sequencing. Suspected mutations were verified by excluding possible single nucleotide polymorphisms and bioinformatics analysis. Transcripts of the EXT1 gene in the proband were analyzed by TA clone-sequencing, with its abundance compared with that of healthy controls.

RESULTSDNA sequencing has identified in the proband a novel heterozygous point mutation (c.1164+1G to A) at the 5'splice sites of intron 3 of the EXT1 gene. The same mutation was not found in the healthy controls. Bioinformatics analysis indicated that the mutation is highly conserved and can lead to skipping of exon 3 or aberrant splicing. TA clone-sequencing indicated that the numbers of transcripts with skipping of exon 3 has significantly increased in the proband (< 0.05) compared with the controls.

CONCLUSIONThe c.1164+1G to A mutation has resulted in skipping of exon 3 in a proportion of EXT1 gene transcripts. As the result, the number of transcripts with tumor suppressing function is relatively reduced and has ultimately led to the tumors.

Adult ; Base Sequence ; Child ; Exostoses, Multiple Hereditary ; genetics ; Female ; Humans ; Male ; Molecular Sequence Data ; N-Acetylglucosaminyltransferases ; genetics ; Point Mutation ; RNA Splice Sites ; RNA Splicing

OBJECTIVETo analyze mutations of SLC26A4 gene and explore their origins for a patient with enlarge vestibuar aqueduct syndrome.

METHODSClinical data and peripheral venous blood samples were collected from the patient and her parents. Genome DNA was extracted from the peripheral blood. All of the 21 exons of the SLC26A4 gene were amplified with PCR and subjected to directly sequencing.

RESULTSThe patient was found to have carried two mutant alleles of the SLC26A4 gene, namely c.1522A to G and c.1229C to T, which were inherited from her father and mother, respectively.

CONCLUSIONSLC26A4 c.1522A to G is likely to be a pathogenic mutation. Above results may facilitate genetic counseling and prenatal diagnosis for this family.

Adult ; Amino Acid Sequence ; Child ; Exons ; Female ; Hearing Loss, Sensorineural ; genetics ; Humans ; Male ; Membrane Transport Proteins ; genetics ; Molecular Sequence Data ; Pedigree ; Vestibular Aqueduct ; abnormalities

OBJECTIVETo detect pathogenic mutation of the SLC39A4 gene in a male patient with acrodermatitis enteropathica (AE).

METHODSPeripheral venous blood sample and clinical data from the patient and his parents were collected. One hundred unrelated healthy individuals were recruited as controls. All coding exons and flanking exon-intron sequences of the SLC39A4 gene were analyzed by PCR and direct sequencing.

RESULTSThe results revealed that the patient and his mother have both carried a novel frame-shift mutation c.1110InsG (p.Gly370GlyfsX47 to TGA) in exon 6. A novel nonsense mutation c.958C to T (p.Q320X) in exon 5 was also detected in the patient and his father and grandmother. This novel mutation was not detected in the unaffected family members and 100 unrelated healthy controls.

CONCLUSIONThe novel frame-shift mutation c.1110InsG (p.Gly370GlyfsX47 to TGA) derived from the mother and nonsense mutation c.958C to T (p.Q320X) of the SLC39A4 gene derived from the father may underlie the disease in the patient.

Acrodermatitis ; genetics ; Adolescent ; Base Sequence ; Cation Transport Proteins ; genetics ; Exons ; Homozygote ; Humans ; Male ; Molecular Sequence Data ; Mutation ; Pedigree ; Zinc ; deficiency

OBJECTIVETo delineate the clinical, biochemical and genetic mutational characteristics of a child with mitochondrial complex III deficiency.

METHODSClinical information and results of auxiliary examination of the patient were analyzed. Next-generation sequencing of the mitochondrial genome and related nuclear genes was carried out. Suspected mutation was confirmed in both parents with Sanger sequencing. Heterozygous deletion was mapped with chromosomal microarray analysis and confirmed with real-time PCR.

RESULTSThe patient presented with vomiting, polypnea, fever, metabolic acidosis, hyperlactatemia, hypoglycemia, dysfunction of coagulation and immune system, in addition with increased lactate dehydrogenase and creatine kinase isoenzyme. Elevation of blood alanine and acylcarnitines as well as urinary ketotic dicarboxylic acid were also noted. The patient also presented development delay, mental retardation and hypotonia. Sequence analysis revealed two mutations in the nuclear gene UQCRB, which included a previously reported frameshift mutation c.306_309delAAAA(p.Arg105Lysfs*22) and a novel large deletion encompassing the entire UQCRB gene.

CONCLUSIONThe clinical, biochemical and gene mutation characteristics of a child with mitochondrial complex III deficiency caused by mutations of the UQCRB gene have been delineated.

Adult ; Base Sequence ; Carrier Proteins ; genetics ; Electron Transport Complex III ; deficiency ; genetics ; Female ; Humans ; Infant ; Male ; Mitochondrial Diseases ; genetics ; Molecular Sequence Data ; Mutation

OBJECTIVETo detect potential mutation of the TCIRG1 gene in a boy with infantile malignant osteopetrosis.

METHODSTarget sequence capture and next-generation sequencing were applied for the proband and his parents to identify the causative mutation, and Sanger sequencing was used to verify the suspected mutation.

RESULTSThe proband manifested at 4 months of age with symptoms including anemia, thrombocytopenia, hepatosplenomegaly, and cephalus quadratus. X-ray revealed generalized increased bone density. A novel compound heterozygous mutation, c.796G to T (p.E266X) and c.1372G to A (p.G458S), were identified in the boy. His father and grandmother also carried the c.796G to T (p.E266X) mutation, and his mother carried the c.1372G to A (p.G458S) mutation. Neither mutation was found in the PubMed and ClinVar databases.

CONCLUSIONThe novel compound heterozygous mutation c.796G to T (p.E266X) and c.1372G to A (p.G458S) probably underlies the disease in the proband. Above results may enrich the mutation spectrum of the TCIRG1 gene and provide new evidence for the molecular basis of infantile malignant osteopetrosis.

Adult ; Asian Continental Ancestry Group ; Base Sequence ; Humans ; Infant ; Infant, Newborn ; Infant, Newborn, Diseases ; genetics ; Male ; Middle Aged ; Molecular Sequence Data ; Mutation ; Osteopetrosis ; genetics ; Pedigree ; Vacuolar Proton-Translocating ATPases ; genetics

OBJECTIVETo analyze the clinicopathologic features and genetic mutation in a patient diagnosed with focal segmental glomerulosclerosis (FSGS).

METHODSClinicopathologic data of the patient, who was diagnosed with primary FSGS by renal biopsy, was collected. Mutations of FSGS-related genes were screened with next-generation sequencing. Suspected pathogenic mutation was verified with Sanger sequencing.

RESULTSNext-generation sequencing detected a missense mutation (c.2215C to G, p.P739A) in exon 28 of the COL4A5 gene in the patient. The same mutation was also detected in his mother who was asymptomatic. Another missense mutation (c.2215C to T, p.P739S) in the same codon has been related with Alport syndrome.

CONCLUSIONThe c.2215C to G (p.P739A) mutation may be one of pathogenic mutations underlying FSGS. This has provided further evidence for the phenotypic heterogeneity of COL4A5 gene mutations.

Adult ; Base Sequence ; Collagen Type IV ; genetics ; Exons ; Female ; Glomerulosclerosis, Focal Segmental ; genetics ; High-Throughput Nucleotide Sequencing ; Humans ; Male ; Molecular Sequence Data ; Mutation ; Young Adult