[Objective] The sequences of mitochondrial DNA D-loop region of Xinjiang Goose with three different colors of plumage were analyzed in order to study the genetic diversity of Xinjiang Goose, as well as the phylogeny a...[Objective] The sequences of mitochondrial DNA D-loop region of Xinjiang Goose with three different colors of plumage were analyzed in order to study the genetic diversity of Xinjiang Goose, as well as the phylogeny and evolution. [Method] Ten geese were selected randomly from the core populations of grey-, mosaic- and white-plumaged Xinjiang Goose respectively with a total number of thirty as experi- mental materials, of which the blood samples were collected from the largest vein under the wing （brachial vein） for DNA extraction. Sequences of mitochondrial DNA D-loop regions were determined using DNA sequencing technology to analyze the polymorphism. In addition, the genetic distances among different populations were estimated through the comparison with the reference sequences. [Resull] The con- tents of A, G, C and T nucleotides in the D-loop region of Xinjiang Goose were 28.85%, 17.05%, 25.38% and 28.72%, respectively. The average haplotype diversity and nucleotide diversity of Xinjiang Goose were 0.583 and 0.056. Xinjiang Goose and Greylag Goose were clustered into the same group. [Conclusion] The results showed that Xinjiang Geese with three different colors of plumage all descend from Greylag Goose （Anser anser）.展开更多
Telomeric repeat amplification protocol (TRAP) is now a conventional assay for detecting telomerase activity. However, this method presents problems owing to tedious quantitation, radioisotopic handling. In order to a...Telomeric repeat amplification protocol (TRAP) is now a conventional assay for detecting telomerase activity. However, this method presents problems owing to tedious quantitation, radioisotopic handling. In order to alleviate these inconveniences, a novel telomerase DNA sequencing assay together with TRAP to detect human telomerase activity was developed. It was used to detect telomerase activity in Hela, HLF, MCF, K562, SMMC 7721 cells, Leukocytes and RNase pretreated or heat treated cells as control. Telomerase activity assayed by this method was positive when the number of K562 cells examined was 102,103, and 104. The telomerase activity depended on the number of K562 cells used in the assay. Telomerase activity of Rnase pretreated cells or heat treated cells, and human normal peripheral blood leukocyte(Leu) were negative. The result of this method was available within a few hours and was handled without radioisotope. Further studies should be taken to detect telomerase activity in quantitation.展开更多
A new genomic DNA encoding a member of Gastrodia antifungal protein family is isolated and sequenced. This gene contains a 510 bp open reading frame and 531 bp promoter region without introns. Sequence analysis indica...A new genomic DNA encoding a member of Gastrodia antifungal protein family is isolated and sequenced. This gene contains a 510 bp open reading frame and 531 bp promoter region without introns. Sequence analysis indicates that a 28 amino acids signal peptide exists at the N terminal. It shows high sequence homology with the mannose binding lectins from Epipactis helleborine, Listera ovata and Cymbidium hybrid. A putative TATA box and transcription start site is detected in the promoter region.展开更多
A technique of direct sequence analysis of β-globin gene with the products of amplifi-cation by polymerase chain reaction (PCR) was reported and a case of β-thalassemia with therare mutation in Chinese,‘codon 14/15...A technique of direct sequence analysis of β-globin gene with the products of amplifi-cation by polymerase chain reaction (PCR) was reported and a case of β-thalassemia with therare mutation in Chinese,‘codon 14/15 (+G)’ was detected by this method.After the se-quence of the mutation site was determined,an analysis of the restriction map of the gene anddot blot hybridization with radioactive allele specific oligonucleotide probe was designed to con-firm the result of DNA sequencing.展开更多
Objective:To demonstrate DNA sequencing analysis (DNAsa) of sinus cultures in patients with CRS is a reliable method of detecting pathogens in polymicrobial CRS infections.Methods:After obtaining Institutional Review ...Objective:To demonstrate DNA sequencing analysis (DNAsa) of sinus cultures in patients with CRS is a reliable method of detecting pathogens in polymicrobial CRS infections.Methods:After obtaining Institutional Review Board approval for this prospective cohort study,we selected a random sample of 50 patients with CRS at Medstar Georgetown University Hospital between September 2016 and March 2017.We defined CRS as a history of rhinosinusitis refractory to maximal medical therapy and prior endoscopic sinus surgery.Patients demonstrating active purulence in a sinus cavity were prospectively selected to undergo standard hospital cultures (SHC) and DNAsa cultures.Organisms identified in both methods were compared for each patient.Results:Specimens were obtained from 29 female and 16 male patients with a mean age of 50 years.A total of 45 cultures were included in our final analysis;five cultures were excluded after inappropriate laboratory processing.Results from these patients were compared and analyzed.Cohen's weighted kappa analysis showed agreement between the two testing methods in identifying predominant microorganisms.DNAsa detected 31.9% more microorganisms compared to SHC (P < 0.05).When multiple microorganisms were detected,DNAsa yielded more positive results compared to SHC (P < 0.05).Conclusions:DNAsa detects all microorganisms identified by SHC as well as predominant microorganisms not detected by SHC.Thus molecular pathogen identification may be more reliable for identifying multiple microorganisms as compared to standard culture techniques that identify only one or two microorganisms.In recalcitrant cases of CRS,DNAsa may provide better guidance in selection of appropriate antimicrobial treatment.展开更多
While M13mpl8 double-stranded DNA was irradiated with ion beam, and transfected into E. coli JM103, a decrease of transfecting activity was discovered. The lacZ-mutation frequency at 20% survival could reach (3.6-16.8...While M13mpl8 double-stranded DNA was irradiated with ion beam, and transfected into E. coli JM103, a decrease of transfecting activity was discovered. The lacZ-mutation frequency at 20% survival could reach (3.6-16.8) × 104, about 2.3-10 times that of unirradiated M13DNA. Altogether, 27 lacZ~ mutants were select-ed, 10 of which were used for sequencing. 7 of the sequenced mutants show base changes in 250-bp region examined (the remaining 3 mutants probably have base changes outside the regions sequenced). 5 of the base-changed mutants contain more than one mutational base sites (some of them even have 5-6 mutational base sites in 250-bp region ex-amined) ; this dense distribution of base changes in polysites has seldom been seen in X-rays, γ-rays or UV induced DNA mutations. Our experiments also showed that the types of base changes include transitions( 50 % ), transversions (45% ) and deletion (5% ); no addition or duplication was observed. The transitions were mainly C→T and A→G; the transversions were mainly C→A and C→G. The mutations involving cytosine residue (in the template strand) con-stitute about 60% of all the base changes observed. In comparison with the surrounding sequences of mutational base sites, the base located between TG and CT is found to be easily substituted.展开更多
[Objective] The research aimed to construct the discriminant classification model of DNA sequence by combining with the biology knowledge and the mathematical method.[Method] According to the polarity nature of side c...[Objective] The research aimed to construct the discriminant classification model of DNA sequence by combining with the biology knowledge and the mathematical method.[Method] According to the polarity nature of side chain radical in the amino acid,the classification information of amino acid which represented the sequence characteristic from the content and array situation of base was extracted from the different sequences that the amino acid content was different.The four-dimension vector was used to represent.Mahalanobis distance and Fisher discriminant methods were used to classify the given sequence.[Result] In the model,the back substitution rates of sample obtained by two kinds of classification methods were both 100%,and the consistent rate of classification was 90%.[Conclusion] In the model,the calculation method was simple,and the accuracy of classification result was higher.It was superior to the discriminant classification model which was only based on the base content.展开更多
Equine infectious anemia virus strain L (EIAV-L) is the parental virulent virus of equine infectious anemia donkey leukocyte attenuated vaccine (DLA EIAV). In this study, peripheral blood leukocytes(PBL) were collecte...Equine infectious anemia virus strain L (EIAV-L) is the parental virulent virus of equine infectious anemia donkey leukocyte attenuated vaccine (DLA EIAV). In this study, peripheral blood leukocytes(PBL) were collected from a horse infected with EIAV-L.The PBL DNAs were extracted.The EIAV-L proviral DNA was amplified in four parts covering the entire proviral genomic sequence by polymerase chain reaction (PCR). Each of the four parts was cloned into the plasmid pBluescript SK, and the recombinant plasmids were designated as p2.8, p2.4. p3.1, and p1.2 respectively. After identification with restriction digestion, the inserts within the four plasmids were sequenced. The complete nucleotide sequence of EIAV-L provirus was determined by analyzing each of the four parts and connecting them as a whole. The genome of EIAV-L is 8235 bp in length, and G + C content is 38%. The comparison analysis by the computer software DNASIS showed that the sequence of EIAV-L shares 98.4% and 96.9% identities with that of D-A EIAV and DLA EIAV respectively. The high homology between these strains showed that they were genetically related. The homology between EIAV-L and D-A EIAV is higher than that between EIAV-L and DLA EIAV, and this is consistent with the derivation progress of DLA EIAV. At both ends of EIAV-L provirus, there is an identical long terminal repeat (LTR) sequence of 316bp in length. The LTR consists of U3, R, and U5 regions. The genome of EIAV-L provirus has three long open reading frames(ORF) corresponding to gag, pol and env genes respectively. The gag gene is 1200bp and located at position 613-1912nt. The pol gene is 3402bp and located at position 1708-5109nt. There is a termination codon within the env dividing it into two parts, envl of 699bp (position 5305-6003nt)and env2 of 1827bp (position 6073-7899nt). The provirus has three additional small ORFs: S1, S2 and S3 with sizes of 153bp(position 5113-5265nt), 204bp(position 5279-5482nt)and 402bp(position 7245-7646nt) respectively.展开更多
Objective: To isolate human tumor metastasis suppressive DNA sequence and to study the molecular mechanisms regulating tumor metastasis. Methods: A mouse lung adenocarcinoma cell clone 12 derived from its parent cell ...Objective: To isolate human tumor metastasis suppressive DNA sequence and to study the molecular mechanisms regulating tumor metastasis. Methods: A mouse lung adenocarcinoma cell clone 12 derived from its parent cell line LM2, which had been transduced with normal human genomic DNA, was previously reported. Compared with LM2, the metastatic potential of clone 12 was very much decreased. Clone 12 was used in this study to amplify the human DNA fragments by Inter Alu PCR technique. The human DNA fragments obtained were then transfected into LM2 cells and their malignant phenotype was tested in vitro and in vivo, and compared with that of the untransfected LM2 cells.Results Three human DNA fragments of 700, 500 and 300 bp were isolated. DNA sequencing revealed that the 700bp fragment does not show homology with hitherto reported genes and was accepted by the Genbank (pt712 U67835). In vitro proliferation and colony formation in soft agar of the 700 bp fragment-transfected LM2 cells were significantly inhibited as compared to the untransfected LM2 cells. Upon subcutaneous inoculation to syngeneic T739 mice, the 700bp-transfected LM2 cells grew more slowly and smaller tumors developed compared to the untransfected ones. Moreover, lung metastasis was not found in 6 of 10 mice inoculated with the 700bp-transfected LM2 cells, while it was found in 9 of 10 mice inoculated with the untransfected LM2 cells. The difference was statistically significant (P<0.001). The frequency of lymph node metastasis was also statistically different between the 2 groups of mice.Conclusion The newly isolated 700bp human DNA fragment may be a metastasis suppressor gene of malignant tumor.展开更多
A new method is presented with which we isolated milochondrial DNA from fresh carp liver usingdifferential centrifugation and DNase treatment that gave high yield of purified product with an easyand economical procedu...A new method is presented with which we isolated milochondrial DNA from fresh carp liver usingdifferential centrifugation and DNase treatment that gave high yield of purified product with an easyand economical procedure. Highly distinct bands were displayed in agarose gel electrophoresls ofthe product digested with restrictlon enzymes, which were successfully used in constructingrestriction map and molecular clone of mitochondrial genes. With DNAs thus obtained, we havecloned cysteine tRNA gene (tRNA^(Cys) gene) of carp mitochondria, determined the nucleotide sequenceof it and the light strand origin, and depicted the cloverleaf secondary structure of tDNA^(Cya) and thelight strand origin. Analysis of nucleotide sequences of tRNA^(Cy) genes of 5 vertebrates has revealedunusual features of carp mitochondrial tRNA^(Cy) gene as compared with their cytoplasmic counter-parts, Altogether 36 bases were found in the light strand origin of carp mitochondriaf: 11 pairs in thestem; and 14 bases in the loop. As compared with those of other 11 vertebrate species, the sequenceof the stem is very conservative while both sequence and length of the loop are quite variable. Thestructure of the stem-loop may play an important role in light strand replication.展开更多
AIM:To compare the differences between dideoxy sequencing/KRAS StripAssay/pyrosequencing for detection of KRAS mutation in Chinese colorectal cancer (CRC) patients.METHODS:Formalin-f ixed, paraff in-embedded (FFPE) sa...AIM:To compare the differences between dideoxy sequencing/KRAS StripAssay/pyrosequencing for detection of KRAS mutation in Chinese colorectal cancer (CRC) patients.METHODS:Formalin-f ixed, paraff in-embedded (FFPE) samples with tumor cells ≥ 50% were collected from 100 Chinese CRC patients at Beijing Cancer Hospital. After the extraction of genome DNA from FFPE samples, fragments contained codons 12 and 13 of KRAS exon 2 were amplified by polymerase chain reaction and analyzed by dideoxy sequencing, the KRAS Strip Assay and pyrosequencing. In addition, the sensitivities of the 3 methods were compared on serial dilutions (contents of mutant DNA: 100%,50%,20%, 5%,10%, 5%,1%,0%) of A549 cell line DNA (carrying the codon 12 Gly>Ser mutation) into wild-type DNA (human normal intestinal mucosa). The results of dideoxy sequencing,the KRAS StripAssay and pyrosequencing were analyzed by Chromas Software, Collector forKRAS Strip Assay and the pyrosequencing PyroMarkTM Q24 system, respectively.RESULTS: Among 100 patients, KRAS mutations were identif ied in 34%, 37% and 37% of patients by dideoxy sequencing, the KRAS StripAssay and pyrosequencing, respectively. The sensitivity was highest with the KRAS Strip Assay (1%), followed by pyrosequencing (5%), and dideoxy sequencing was lowest (15%). Six different mutation types were found in this study with 3 main mutations Gly12 Asp (GGT>GAT), Gly12 Val (GGT>GTT) and Gly13 Asp (GGC>GAC). Thirty-three patients were identifi ed to have KRAS mutations by the 3 methods, and a total of 8 patients had conflicting results between 3 methods: 4 mutations not detected by dideoxy sequencing and the KRAS StripAssay were identified by pyrosequencing; 3 mutations not detected by dideoxy sequencing and pyrosequencing were identif ied by the KRAS StripAssay; and 1 mutation not detected by pyrosequencing was conf irmed by dideoxy sequencing and the KRAS StripAssay. Among these discordant results, the results identif ied by dideoxy sequencing were consistent either with the KRAS StripAssay or with pyrosequencing, which indicated that the accuracy of dideoxy sequencing was high. CONCLUSION: Taking a worldwide view of reports and our results,dideoxy sequencing remains the most popular method because of its low cost and high accuracy.展开更多
MtDNA was successfully extracted from ten individual bones (femurs) in the tombs of ancient Jushi in Turfan basin, dated back to the year about 3 000-2 500 years ago. By means of four overlapping primers, we got nucl...MtDNA was successfully extracted from ten individual bones (femurs) in the tombs of ancient Jushi in Turfan basin, dated back to the year about 3 000-2 500 years ago. By means of four overlapping primers, we got nucleotide sequence of the 218bp length. Ancient mtDNA was analyzed by the sequencing of hypervariable region Ⅰ of the mtDNA control region. The result shows that 9 haplotypes with 24 polymorphic sites were obtained. The phylogenetic analysis indicated that Mongolians and Altai are the population genetically closest to the Jushi groups and Jushi mtDNA pool being an admixture of eastern Asian and European lineages. So our preliminary data imply that an ancient mingling of Euro-Asian population had existed in Turfan basin prior to the early Iron Age.展开更多
Stream cipher, DNA cryptography and DNA analysis are the most important R&D fields in both Cryptography and Bioinformatics. HC-256 is an emerged scheme as the new generation of stream ciphers for advanced network ...Stream cipher, DNA cryptography and DNA analysis are the most important R&D fields in both Cryptography and Bioinformatics. HC-256 is an emerged scheme as the new generation of stream ciphers for advanced network security. From a random sequencing viewpoint, both sequences of HC-256 and real DNA data may have intrinsic pseudo-random properties respectively. In a recent decade, many DNA sequencing projects are developed on cells, plants and animals over the world into huge DNA databases. Researchers notice that mammalian genomes encode thousands of large noncoding RNAs (lncRNAs), interact with chromatin regulatory complexes, and are thought to play a role in localizing these complexes to target loci across the genome. It is a challenge target using higher dimensional visualization tools to organize various complex interactive properties as visual maps. The Variant Map System (VMS) as an emerging scheme is systematically proposed in this paper to apply multiple maps that used four Meta symbols as same as DNA or RNA representations. System architecture of key components and core mechanism on the VMS are described. Key modules, equations and their I/O parameters are discussed. Applying the VM System, two sets of real DNA sequences from both sample human (noncoding DNA) and corn (coding DNA) genomes are collected in comparison with pseudo DNA sequences generated by HC-256 to show their intrinsic properties in higher levels of similar relationships among relevant DNA sequences on 2D maps. Sample 2D maps are listed and their characteristics are illustrated under controllable environment. Visual results are briefly analyzed to explore their intrinsic properties on selected genome sequences.展开更多
A new version of DNA walks, where nucleotides are regarded unequal in their contribution to a walk is introduced, which allows us to study thoroughly the “fine structure” of nucleotide sequences. The approach is bas...A new version of DNA walks, where nucleotides are regarded unequal in their contribution to a walk is introduced, which allows us to study thoroughly the “fine structure” of nucleotide sequences. The approach is based on the assumption that nucleotides have an inner abstract characteristic, the determinative degree, which reflects genetic code phenomenological prop-erties and is adjusted to nucleotides physical properties. We consider each codon position independently, which gives three separate walks characterized by different angles and lengths, and that such an object is called triander which reflects the “strength” of branch. A general method for identifying DNA sequence “by triander” which can be treated as a unique “genogram” (or “gene passport”) is proposed. The two- and three-dimensional trianders are considered. The difference of sequences fine structure in genes and the intergenic space is shown. A clear triplet signal in coding sequences was found which is absent in the intergenic space and is independent from the sequence length. This paper presents the topological classification of trianders which can allow us to provide a detailed working out signatures of functionally different genomic regions.展开更多
In a recent decade, many DNA sequencing projects are developed on cells, plants and animals over the world into huge DNA databases. Researchers notice that mammalian genomes encoding thousands of large noncoding RNAs ...In a recent decade, many DNA sequencing projects are developed on cells, plants and animals over the world into huge DNA databases. Researchers notice that mammalian genomes encoding thousands of large noncoding RNAs (lncRNAs), interact with chromatin regulatory complexes, and are thought to play a role in localizing these complexes to target loci across the genome. It is a challenge target using higher dimensional tools to organize various complex interactive properties as visual maps. In this paper, a Pseudo DNA Variant MapPDVM is proposed following Cellular Automata to represent multiple maps that use four Meta symbols as well as DNA or RNA representations. The system architecture of key components and the core mechanism on the PDVM are described. Key modules, equations and their I/O parameters are discussed. Applying the PDVM, two sets of real DNA sequences from both the sample human (noncoding DNA) and corn (coding DNA) genomes are collected in comparison with two sets of pseudo DNA sequences generated by a stream cipher HC-256 under different modes to show their intrinsic properties in higher levels of similar relationships among relevant DNA sequences on 2D maps. Sample 2D maps are listed and their characteristics are illustrated under a controllable environment. Various distributions can be observed on both noncoding and coding conditions from their symmetric properties on 2D maps.展开更多
AIM To explore a rapid and easy sequencing method for hepatitis C virus (HCV) genome, and establish a new sequencing method in China. METHODS Polymerase Chain Reaction (PCR) was combined with DNA sequencing techn...AIM To explore a rapid and easy sequencing method for hepatitis C virus (HCV) genome, and establish a new sequencing method in China. METHODS Polymerase Chain Reaction (PCR) was combined with DNA sequencing technique. PCR products were purified by agarose gel electrophoresis, polyacrylamide gel electrophoresis (PAGE), Polyethylene glycol (PEG) respectively. Then in the presence of a 5′ labeling PCR primer, purified PCR products were directly sequenced. By this method, HCV NS5b cDNA from two HCV infected individuals (HC 42 and HC 49) were sequenced.展开更多
Since the 1980s,Sargassum fusiforme has been cultivated in Zhejiang,South China,and nowadays it becomes one of the important commercial seaweeds in China.With traditions of eating habits in the East Asian countries,th...Since the 1980s,Sargassum fusiforme has been cultivated in Zhejiang,South China,and nowadays it becomes one of the important commercial seaweeds in China.With traditions of eating habits in the East Asian countries,this brown alga is used as food,because it contained functional oligo/polysaccharides and chemical components,and was regarded playing roles in antioxidant activities and regulating immunology.Through over 15 years’selection,breeding and cultivation,we obtained three strains with good traits and testified their characters during the production,which included the cultivars with high yield and other two good characters,either all the selected strains were applied in the Sargassum production.To avoid confusion during the selection and nursery,it was preferred to establish one fingerprint for distinguishing the Sargassum cultivars from different strains.Random amplified polymorphic DNA(RAPD)and inter-simple sequence repeat(ISSR)methods were adopted to analyze the genetic diversities of the selected S.fusiforme strains.With that,one fingerprint with RAPD markers was constructed,and one sequence characterized amplifi ed region(SCAR)marker to S.fusiforme was obtained.It is indicated that the applied fingerprint could be valid in S.fusiforme genetic and germplasm justification,and will be positive to molecular marker assistance in its selection and cultivation.展开更多
AIM: To study the clinicopathological and molecular genetic characteristics of typical Chinese hereditary nonpolyposis cotorectal cancer (HNPCC) families. METHODS: Four typical Chinese HNPCC families were analyzed usi...AIM: To study the clinicopathological and molecular genetic characteristics of typical Chinese hereditary nonpolyposis cotorectal cancer (HNPCC) families. METHODS: Four typical Chinese HNPCC families were analyzed using microdissection, microsatellite instability analysis, immunostaining of hMSH2 and hMLH1 proteins and direct DNA sequencing of hMSH2 and hMLH1 genes. RESULTS: All five tumor tissues of 4 probands from the 4 typical Chinese HNPCC families showed microsatellite instability at more than two loci (MSI-H or RER+ phenotype). Three out of the 4 cases lost hMSH2 protein expression and the other case showed no hMLH1 protein expression. Three pathological germline mutations (2 in hMSH2 and 1 in hMLH1), which had not been reported previously, were identified. The same mutations were also found in other affected members of two HNPCC families,respectively. CONCLUSION: Typical Chinese HNPCC families showed relatively frequent germline mutation of mismatch repair genes. High-level microsatellite instability and loss of expression of mismatch repair genes correlated closely with germline mutation of mismatch repair genes. Microsatellite instability analysis and immunostaining of mismatch repair gene might serve as effective screening methods before direct DNA sequencing. It is necessary to establish clinical criteria and molecular diagnostic strategies more suitable for Chinese HNPCC families.展开更多
AIM:To isolate and analyze the DNA sequences which are methylated differentially between gastric cancer and normal gastric mucosa. METHODS: The differentially methylated DNA sequences between gastric cancer and normal...AIM:To isolate and analyze the DNA sequences which are methylated differentially between gastric cancer and normal gastric mucosa. METHODS: The differentially methylated DNA sequences between gastric cancer and normal gastric mucosa were isolated by methylation-sensitive representational difference analysis (MS-RDA). Similarities between the separated fragments and the human genomic DNA were analyzed with Basic Local Alignment Search Tool (BLAST). RESULTS: Three differentially methylated DNA sequences were obtained, two of which have been accepted by GenBank. The accession numbers are AY887106 and AY887107. AY887107 was highly similar to the 11th exon of LOC440683 (98%), 3' end of LOC440887 (99%), and promoter and exon regions of DRD5 (94%). AY887106 was consistent (98%) with a CpG island in ribosomal RNA isolated from colorectal cancer by Minoru Toyota in 1999. CONCLUSION: The methylation degree is different between gastric cancer and normal gastric mucosa. The differentially methylated DNA sequences can be isolated effectively by MS-RDA.展开更多
Diversity analysis among 23 rice varieties including 16 non-basmati scented accessions, 5 basmati accessions and 2 non-scented accessions was performed by random amplified polymorphic DNA （RAPD） and inter-simple seq...Diversity analysis among 23 rice varieties including 16 non-basmati scented accessions, 5 basmati accessions and 2 non-scented accessions was performed by random amplified polymorphic DNA （RAPD） and inter-simple sequence repeat （ISSR） marker systems. The varieties analyzed by 11 RAPD and 8 ISSR primers yielded an average of 65% and 80% polymorphism, respectively. The average number of polymorphic bands generated per RAPD primer was 6 and per ISSR primer was 5.87. RAPD and ISSR data analysis individually could not segregate basmati and non-basmati scented rice accessions. However, the analysis using a combined data could group basmati and non-basmati scented rice accessions separately. The bands present specifically among three accessions of non-basmati scented rice were also identified. The study revealed a high genetic diversity among non-basmati scented rice accessions.展开更多
基金Supported by the Fond for Open Projects of Xinjiang Key Laboratory of Herbivore Nutrition for Meat&Milk Production~~
文摘[Objective] The sequences of mitochondrial DNA D-loop region of Xinjiang Goose with three different colors of plumage were analyzed in order to study the genetic diversity of Xinjiang Goose, as well as the phylogeny and evolution. [Method] Ten geese were selected randomly from the core populations of grey-, mosaic- and white-plumaged Xinjiang Goose respectively with a total number of thirty as experi- mental materials, of which the blood samples were collected from the largest vein under the wing （brachial vein） for DNA extraction. Sequences of mitochondrial DNA D-loop regions were determined using DNA sequencing technology to analyze the polymorphism. In addition, the genetic distances among different populations were estimated through the comparison with the reference sequences. [Resull] The con- tents of A, G, C and T nucleotides in the D-loop region of Xinjiang Goose were 28.85%, 17.05%, 25.38% and 28.72%, respectively. The average haplotype diversity and nucleotide diversity of Xinjiang Goose were 0.583 and 0.056. Xinjiang Goose and Greylag Goose were clustered into the same group. [Conclusion] The results showed that Xinjiang Geese with three different colors of plumage all descend from Greylag Goose （Anser anser）.
基金LiaoningScience&TechnologyPlanFoundation No :993 0 5 0 0 1
文摘Telomeric repeat amplification protocol (TRAP) is now a conventional assay for detecting telomerase activity. However, this method presents problems owing to tedious quantitation, radioisotopic handling. In order to alleviate these inconveniences, a novel telomerase DNA sequencing assay together with TRAP to detect human telomerase activity was developed. It was used to detect telomerase activity in Hela, HLF, MCF, K562, SMMC 7721 cells, Leukocytes and RNase pretreated or heat treated cells as control. Telomerase activity assayed by this method was positive when the number of K562 cells examined was 102,103, and 104. The telomerase activity depended on the number of K562 cells used in the assay. Telomerase activity of Rnase pretreated cells or heat treated cells, and human normal peripheral blood leukocyte(Leu) were negative. The result of this method was available within a few hours and was handled without radioisotope. Further studies should be taken to detect telomerase activity in quantitation.
文摘A new genomic DNA encoding a member of Gastrodia antifungal protein family is isolated and sequenced. This gene contains a 510 bp open reading frame and 531 bp promoter region without introns. Sequence analysis indicates that a 28 amino acids signal peptide exists at the N terminal. It shows high sequence homology with the mannose binding lectins from Epipactis helleborine, Listera ovata and Cymbidium hybrid. A putative TATA box and transcription start site is detected in the promoter region.
文摘A technique of direct sequence analysis of β-globin gene with the products of amplifi-cation by polymerase chain reaction (PCR) was reported and a case of β-thalassemia with therare mutation in Chinese,‘codon 14/15 (+G)’ was detected by this method.After the se-quence of the mutation site was determined,an analysis of the restriction map of the gene anddot blot hybridization with radioactive allele specific oligonucleotide probe was designed to con-firm the result of DNA sequencing.
文摘Objective:To demonstrate DNA sequencing analysis (DNAsa) of sinus cultures in patients with CRS is a reliable method of detecting pathogens in polymicrobial CRS infections.Methods:After obtaining Institutional Review Board approval for this prospective cohort study,we selected a random sample of 50 patients with CRS at Medstar Georgetown University Hospital between September 2016 and March 2017.We defined CRS as a history of rhinosinusitis refractory to maximal medical therapy and prior endoscopic sinus surgery.Patients demonstrating active purulence in a sinus cavity were prospectively selected to undergo standard hospital cultures (SHC) and DNAsa cultures.Organisms identified in both methods were compared for each patient.Results:Specimens were obtained from 29 female and 16 male patients with a mean age of 50 years.A total of 45 cultures were included in our final analysis;five cultures were excluded after inappropriate laboratory processing.Results from these patients were compared and analyzed.Cohen's weighted kappa analysis showed agreement between the two testing methods in identifying predominant microorganisms.DNAsa detected 31.9% more microorganisms compared to SHC (P < 0.05).When multiple microorganisms were detected,DNAsa yielded more positive results compared to SHC (P < 0.05).Conclusions:DNAsa detects all microorganisms identified by SHC as well as predominant microorganisms not detected by SHC.Thus molecular pathogen identification may be more reliable for identifying multiple microorganisms as compared to standard culture techniques that identify only one or two microorganisms.In recalcitrant cases of CRS,DNAsa may provide better guidance in selection of appropriate antimicrobial treatment.
文摘While M13mpl8 double-stranded DNA was irradiated with ion beam, and transfected into E. coli JM103, a decrease of transfecting activity was discovered. The lacZ-mutation frequency at 20% survival could reach (3.6-16.8) × 104, about 2.3-10 times that of unirradiated M13DNA. Altogether, 27 lacZ~ mutants were select-ed, 10 of which were used for sequencing. 7 of the sequenced mutants show base changes in 250-bp region examined (the remaining 3 mutants probably have base changes outside the regions sequenced). 5 of the base-changed mutants contain more than one mutational base sites (some of them even have 5-6 mutational base sites in 250-bp region ex-amined) ; this dense distribution of base changes in polysites has seldom been seen in X-rays, γ-rays or UV induced DNA mutations. Our experiments also showed that the types of base changes include transitions( 50 % ), transversions (45% ) and deletion (5% ); no addition or duplication was observed. The transitions were mainly C→T and A→G; the transversions were mainly C→A and C→G. The mutations involving cytosine residue (in the template strand) con-stitute about 60% of all the base changes observed. In comparison with the surrounding sequences of mutational base sites, the base located between TG and CT is found to be easily substituted.
基金Supported by Science Research Project of Ningbo Dahongying University in2011(CF102601)~~
文摘[Objective] The research aimed to construct the discriminant classification model of DNA sequence by combining with the biology knowledge and the mathematical method.[Method] According to the polarity nature of side chain radical in the amino acid,the classification information of amino acid which represented the sequence characteristic from the content and array situation of base was extracted from the different sequences that the amino acid content was different.The four-dimension vector was used to represent.Mahalanobis distance and Fisher discriminant methods were used to classify the given sequence.[Result] In the model,the back substitution rates of sample obtained by two kinds of classification methods were both 100%,and the consistent rate of classification was 90%.[Conclusion] In the model,the calculation method was simple,and the accuracy of classification result was higher.It was superior to the discriminant classification model which was only based on the base content.
基金by National Science Foundation of China(No.39470535).
文摘Equine infectious anemia virus strain L (EIAV-L) is the parental virulent virus of equine infectious anemia donkey leukocyte attenuated vaccine (DLA EIAV). In this study, peripheral blood leukocytes(PBL) were collected from a horse infected with EIAV-L.The PBL DNAs were extracted.The EIAV-L proviral DNA was amplified in four parts covering the entire proviral genomic sequence by polymerase chain reaction (PCR). Each of the four parts was cloned into the plasmid pBluescript SK, and the recombinant plasmids were designated as p2.8, p2.4. p3.1, and p1.2 respectively. After identification with restriction digestion, the inserts within the four plasmids were sequenced. The complete nucleotide sequence of EIAV-L provirus was determined by analyzing each of the four parts and connecting them as a whole. The genome of EIAV-L is 8235 bp in length, and G + C content is 38%. The comparison analysis by the computer software DNASIS showed that the sequence of EIAV-L shares 98.4% and 96.9% identities with that of D-A EIAV and DLA EIAV respectively. The high homology between these strains showed that they were genetically related. The homology between EIAV-L and D-A EIAV is higher than that between EIAV-L and DLA EIAV, and this is consistent with the derivation progress of DLA EIAV. At both ends of EIAV-L provirus, there is an identical long terminal repeat (LTR) sequence of 316bp in length. The LTR consists of U3, R, and U5 regions. The genome of EIAV-L provirus has three long open reading frames(ORF) corresponding to gag, pol and env genes respectively. The gag gene is 1200bp and located at position 613-1912nt. The pol gene is 3402bp and located at position 1708-5109nt. There is a termination codon within the env dividing it into two parts, envl of 699bp (position 5305-6003nt)and env2 of 1827bp (position 6073-7899nt). The provirus has three additional small ORFs: S1, S2 and S3 with sizes of 153bp(position 5113-5265nt), 204bp(position 5279-5482nt)and 402bp(position 7245-7646nt) respectively.
基金the National Natural Science Foundation of China (No. 39370761 ).
文摘Objective: To isolate human tumor metastasis suppressive DNA sequence and to study the molecular mechanisms regulating tumor metastasis. Methods: A mouse lung adenocarcinoma cell clone 12 derived from its parent cell line LM2, which had been transduced with normal human genomic DNA, was previously reported. Compared with LM2, the metastatic potential of clone 12 was very much decreased. Clone 12 was used in this study to amplify the human DNA fragments by Inter Alu PCR technique. The human DNA fragments obtained were then transfected into LM2 cells and their malignant phenotype was tested in vitro and in vivo, and compared with that of the untransfected LM2 cells.Results Three human DNA fragments of 700, 500 and 300 bp were isolated. DNA sequencing revealed that the 700bp fragment does not show homology with hitherto reported genes and was accepted by the Genbank (pt712 U67835). In vitro proliferation and colony formation in soft agar of the 700 bp fragment-transfected LM2 cells were significantly inhibited as compared to the untransfected LM2 cells. Upon subcutaneous inoculation to syngeneic T739 mice, the 700bp-transfected LM2 cells grew more slowly and smaller tumors developed compared to the untransfected ones. Moreover, lung metastasis was not found in 6 of 10 mice inoculated with the 700bp-transfected LM2 cells, while it was found in 9 of 10 mice inoculated with the untransfected LM2 cells. The difference was statistically significant (P<0.001). The frequency of lymph node metastasis was also statistically different between the 2 groups of mice.Conclusion The newly isolated 700bp human DNA fragment may be a metastasis suppressor gene of malignant tumor.
基金This work was supported by grant from National Foundation of Natural Sciences of China.
文摘A new method is presented with which we isolated milochondrial DNA from fresh carp liver usingdifferential centrifugation and DNase treatment that gave high yield of purified product with an easyand economical procedure. Highly distinct bands were displayed in agarose gel electrophoresls ofthe product digested with restrictlon enzymes, which were successfully used in constructingrestriction map and molecular clone of mitochondrial genes. With DNAs thus obtained, we havecloned cysteine tRNA gene (tRNA^(Cys) gene) of carp mitochondria, determined the nucleotide sequenceof it and the light strand origin, and depicted the cloverleaf secondary structure of tDNA^(Cya) and thelight strand origin. Analysis of nucleotide sequences of tRNA^(Cy) genes of 5 vertebrates has revealedunusual features of carp mitochondrial tRNA^(Cy) gene as compared with their cytoplasmic counter-parts, Altogether 36 bases were found in the light strand origin of carp mitochondriaf: 11 pairs in thestem; and 14 bases in the loop. As compared with those of other 11 vertebrate species, the sequenceof the stem is very conservative while both sequence and length of the loop are quite variable. Thestructure of the stem-loop may play an important role in light strand replication.
文摘AIM:To compare the differences between dideoxy sequencing/KRAS StripAssay/pyrosequencing for detection of KRAS mutation in Chinese colorectal cancer (CRC) patients.METHODS:Formalin-f ixed, paraff in-embedded (FFPE) samples with tumor cells ≥ 50% were collected from 100 Chinese CRC patients at Beijing Cancer Hospital. After the extraction of genome DNA from FFPE samples, fragments contained codons 12 and 13 of KRAS exon 2 were amplified by polymerase chain reaction and analyzed by dideoxy sequencing, the KRAS Strip Assay and pyrosequencing. In addition, the sensitivities of the 3 methods were compared on serial dilutions (contents of mutant DNA: 100%,50%,20%, 5%,10%, 5%,1%,0%) of A549 cell line DNA (carrying the codon 12 Gly>Ser mutation) into wild-type DNA (human normal intestinal mucosa). The results of dideoxy sequencing,the KRAS StripAssay and pyrosequencing were analyzed by Chromas Software, Collector forKRAS Strip Assay and the pyrosequencing PyroMarkTM Q24 system, respectively.RESULTS: Among 100 patients, KRAS mutations were identif ied in 34%, 37% and 37% of patients by dideoxy sequencing, the KRAS StripAssay and pyrosequencing, respectively. The sensitivity was highest with the KRAS Strip Assay (1%), followed by pyrosequencing (5%), and dideoxy sequencing was lowest (15%). Six different mutation types were found in this study with 3 main mutations Gly12 Asp (GGT>GAT), Gly12 Val (GGT>GTT) and Gly13 Asp (GGC>GAC). Thirty-three patients were identifi ed to have KRAS mutations by the 3 methods, and a total of 8 patients had conflicting results between 3 methods: 4 mutations not detected by dideoxy sequencing and the KRAS StripAssay were identified by pyrosequencing; 3 mutations not detected by dideoxy sequencing and pyrosequencing were identif ied by the KRAS StripAssay; and 1 mutation not detected by pyrosequencing was conf irmed by dideoxy sequencing and the KRAS StripAssay. Among these discordant results, the results identif ied by dideoxy sequencing were consistent either with the KRAS StripAssay or with pyrosequencing, which indicated that the accuracy of dideoxy sequencing was high. CONCLUSION: Taking a worldwide view of reports and our results,dideoxy sequencing remains the most popular method because of its low cost and high accuracy.
基金Supported by Fund for the Important Project of the Science and Technology DepartmentMinistry of Education
文摘MtDNA was successfully extracted from ten individual bones (femurs) in the tombs of ancient Jushi in Turfan basin, dated back to the year about 3 000-2 500 years ago. By means of four overlapping primers, we got nucleotide sequence of the 218bp length. Ancient mtDNA was analyzed by the sequencing of hypervariable region Ⅰ of the mtDNA control region. The result shows that 9 haplotypes with 24 polymorphic sites were obtained. The phylogenetic analysis indicated that Mongolians and Altai are the population genetically closest to the Jushi groups and Jushi mtDNA pool being an admixture of eastern Asian and European lineages. So our preliminary data imply that an ancient mingling of Euro-Asian population had existed in Turfan basin prior to the early Iron Age.
文摘Stream cipher, DNA cryptography and DNA analysis are the most important R&D fields in both Cryptography and Bioinformatics. HC-256 is an emerged scheme as the new generation of stream ciphers for advanced network security. From a random sequencing viewpoint, both sequences of HC-256 and real DNA data may have intrinsic pseudo-random properties respectively. In a recent decade, many DNA sequencing projects are developed on cells, plants and animals over the world into huge DNA databases. Researchers notice that mammalian genomes encode thousands of large noncoding RNAs (lncRNAs), interact with chromatin regulatory complexes, and are thought to play a role in localizing these complexes to target loci across the genome. It is a challenge target using higher dimensional visualization tools to organize various complex interactive properties as visual maps. The Variant Map System (VMS) as an emerging scheme is systematically proposed in this paper to apply multiple maps that used four Meta symbols as same as DNA or RNA representations. System architecture of key components and core mechanism on the VMS are described. Key modules, equations and their I/O parameters are discussed. Applying the VM System, two sets of real DNA sequences from both sample human (noncoding DNA) and corn (coding DNA) genomes are collected in comparison with pseudo DNA sequences generated by HC-256 to show their intrinsic properties in higher levels of similar relationships among relevant DNA sequences on 2D maps. Sample 2D maps are listed and their characteristics are illustrated under controllable environment. Visual results are briefly analyzed to explore their intrinsic properties on selected genome sequences.
文摘A new version of DNA walks, where nucleotides are regarded unequal in their contribution to a walk is introduced, which allows us to study thoroughly the “fine structure” of nucleotide sequences. The approach is based on the assumption that nucleotides have an inner abstract characteristic, the determinative degree, which reflects genetic code phenomenological prop-erties and is adjusted to nucleotides physical properties. We consider each codon position independently, which gives three separate walks characterized by different angles and lengths, and that such an object is called triander which reflects the “strength” of branch. A general method for identifying DNA sequence “by triander” which can be treated as a unique “genogram” (or “gene passport”) is proposed. The two- and three-dimensional trianders are considered. The difference of sequences fine structure in genes and the intergenic space is shown. A clear triplet signal in coding sequences was found which is absent in the intergenic space and is independent from the sequence length. This paper presents the topological classification of trianders which can allow us to provide a detailed working out signatures of functionally different genomic regions.
文摘In a recent decade, many DNA sequencing projects are developed on cells, plants and animals over the world into huge DNA databases. Researchers notice that mammalian genomes encoding thousands of large noncoding RNAs (lncRNAs), interact with chromatin regulatory complexes, and are thought to play a role in localizing these complexes to target loci across the genome. It is a challenge target using higher dimensional tools to organize various complex interactive properties as visual maps. In this paper, a Pseudo DNA Variant MapPDVM is proposed following Cellular Automata to represent multiple maps that use four Meta symbols as well as DNA or RNA representations. The system architecture of key components and the core mechanism on the PDVM are described. Key modules, equations and their I/O parameters are discussed. Applying the PDVM, two sets of real DNA sequences from both the sample human (noncoding DNA) and corn (coding DNA) genomes are collected in comparison with two sets of pseudo DNA sequences generated by a stream cipher HC-256 under different modes to show their intrinsic properties in higher levels of similar relationships among relevant DNA sequences on 2D maps. Sample 2D maps are listed and their characteristics are illustrated under a controllable environment. Various distributions can be observed on both noncoding and coding conditions from their symmetric properties on 2D maps.
文摘AIM To explore a rapid and easy sequencing method for hepatitis C virus (HCV) genome, and establish a new sequencing method in China. METHODS Polymerase Chain Reaction (PCR) was combined with DNA sequencing technique. PCR products were purified by agarose gel electrophoresis, polyacrylamide gel electrophoresis (PAGE), Polyethylene glycol (PEG) respectively. Then in the presence of a 5′ labeling PCR primer, purified PCR products were directly sequenced. By this method, HCV NS5b cDNA from two HCV infected individuals (HC 42 and HC 49) were sequenced.
基金Supported by the Doutou Sci-Tech Project(No.N2006Y11B)the Shandong Key Sci-Technology Research Project(Nos.2018SDKJ0302-2,2018SDKJ0502-1)the CAS-Fujian STS Project(No.2017T3012)
文摘Since the 1980s,Sargassum fusiforme has been cultivated in Zhejiang,South China,and nowadays it becomes one of the important commercial seaweeds in China.With traditions of eating habits in the East Asian countries,this brown alga is used as food,because it contained functional oligo/polysaccharides and chemical components,and was regarded playing roles in antioxidant activities and regulating immunology.Through over 15 years’selection,breeding and cultivation,we obtained three strains with good traits and testified their characters during the production,which included the cultivars with high yield and other two good characters,either all the selected strains were applied in the Sargassum production.To avoid confusion during the selection and nursery,it was preferred to establish one fingerprint for distinguishing the Sargassum cultivars from different strains.Random amplified polymorphic DNA(RAPD)and inter-simple sequence repeat(ISSR)methods were adopted to analyze the genetic diversities of the selected S.fusiforme strains.With that,one fingerprint with RAPD markers was constructed,and one sequence characterized amplifi ed region(SCAR)marker to S.fusiforme was obtained.It is indicated that the applied fingerprint could be valid in S.fusiforme genetic and germplasm justification,and will be positive to molecular marker assistance in its selection and cultivation.
基金Supported by the Shanghai Medical Development Project,No.993025
文摘AIM: To study the clinicopathological and molecular genetic characteristics of typical Chinese hereditary nonpolyposis cotorectal cancer (HNPCC) families. METHODS: Four typical Chinese HNPCC families were analyzed using microdissection, microsatellite instability analysis, immunostaining of hMSH2 and hMLH1 proteins and direct DNA sequencing of hMSH2 and hMLH1 genes. RESULTS: All five tumor tissues of 4 probands from the 4 typical Chinese HNPCC families showed microsatellite instability at more than two loci (MSI-H or RER+ phenotype). Three out of the 4 cases lost hMSH2 protein expression and the other case showed no hMLH1 protein expression. Three pathological germline mutations (2 in hMSH2 and 1 in hMLH1), which had not been reported previously, were identified. The same mutations were also found in other affected members of two HNPCC families,respectively. CONCLUSION: Typical Chinese HNPCC families showed relatively frequent germline mutation of mismatch repair genes. High-level microsatellite instability and loss of expression of mismatch repair genes correlated closely with germline mutation of mismatch repair genes. Microsatellite instability analysis and immunostaining of mismatch repair gene might serve as effective screening methods before direct DNA sequencing. It is necessary to establish clinical criteria and molecular diagnostic strategies more suitable for Chinese HNPCC families.
基金The Key Project Fund of the Science and Technology Program of Hunan Province, No. 04SK1004the Scientific Research Fund of Educational Department, Hunan Province, No. 99C195
文摘AIM:To isolate and analyze the DNA sequences which are methylated differentially between gastric cancer and normal gastric mucosa. METHODS: The differentially methylated DNA sequences between gastric cancer and normal gastric mucosa were isolated by methylation-sensitive representational difference analysis (MS-RDA). Similarities between the separated fragments and the human genomic DNA were analyzed with Basic Local Alignment Search Tool (BLAST). RESULTS: Three differentially methylated DNA sequences were obtained, two of which have been accepted by GenBank. The accession numbers are AY887106 and AY887107. AY887107 was highly similar to the 11th exon of LOC440683 (98%), 3' end of LOC440887 (99%), and promoter and exon regions of DRD5 (94%). AY887106 was consistent (98%) with a CpG island in ribosomal RNA isolated from colorectal cancer by Minoru Toyota in 1999. CONCLUSION: The methylation degree is different between gastric cancer and normal gastric mucosa. The differentially methylated DNA sequences can be isolated effectively by MS-RDA.
文摘Diversity analysis among 23 rice varieties including 16 non-basmati scented accessions, 5 basmati accessions and 2 non-scented accessions was performed by random amplified polymorphic DNA （RAPD） and inter-simple sequence repeat （ISSR） marker systems. The varieties analyzed by 11 RAPD and 8 ISSR primers yielded an average of 65% and 80% polymorphism, respectively. The average number of polymorphic bands generated per RAPD primer was 6 and per ISSR primer was 5.87. RAPD and ISSR data analysis individually could not segregate basmati and non-basmati scented rice accessions. However, the analysis using a combined data could group basmati and non-basmati scented rice accessions separately. The bands present specifically among three accessions of non-basmati scented rice were also identified. The study revealed a high genetic diversity among non-basmati scented rice accessions.
