Pseudoalteromonas sp. SM9913 is a phychrotmphic bacterium isolated from the deep-sea sediment. The genes encoding chaperones DnaJ and DnaK of P. sp. SM9913 were cloned by normal PCR and TAIL - PCR (GenBank accession ...Pseudoalteromonas sp. SM9913 is a phychrotmphic bacterium isolated from the deep-sea sediment. The genes encoding chaperones DnaJ and DnaK of P. sp. SM9913 were cloned by normal PCR and TAIL - PCR (GenBank accession Nos DQ640312, DQ504163 ). The chaperones DnaJ and DnaK from the strain SM9913 contain such conserved domains as those of many other bacteria, and show some cold-adapted characteristics in their structures when compared with those from psychro-, meso-and themophilic bacteria. It is indicated that chaperones DnaJ and DnaK of P. sp. SM9913 may be adapted to low temperature in deep-sea and function well in assisting folding, assembling and translocation of proteins at low temperature. This research lays a foundation for the further study on the cold-adapted mechanism of chaperones DnaJ and DnaK of cold-adapted microorganisms.展开更多
The aim of this study was to isolate the full-length cDNA sequences of an unusually highly conserved olfactory receptor (orthologue to the Drosophila melanogaster DOR83b) from the antennae of Spodoptera exigua and S...The aim of this study was to isolate the full-length cDNA sequences of an unusually highly conserved olfactory receptor (orthologue to the Drosophila melanogaster DOR83b) from the antennae of Spodoptera exigua and S. litura by using reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE) methods. Bioinformatics methods were used to further analyze the cDNA sequences and putative amino acid sequences. The two full-length cDNA sequences from olfactory receptor (OR) of male S. exigua and S. litura were named as SexiOR2 and SlitOR2, respectively. SexiOR2 and SlitOR2 consisted of nucleotide sequence of 1 906 and 2 483 bp, respectively, and both with deduced amino acid sequences of 473 residues. The sequence analysis indicated that the deduced amino acid sequences of the cDNA shared the high homologies with OR83b orthologue chemoreceptor sequences from previously reported moths, implying that the cDNA sequences were of OR83b orthologue chemoreceptor genes.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
The sequences of the ITS(internal transcribed spacer) and 5.8S rDNA of three cultivated strains of Porphyra haitanensis thalli(NB,PT and ST) were amplified,sequenced and analyzed.In addition,the phylogenic relationshi...The sequences of the ITS(internal transcribed spacer) and 5.8S rDNA of three cultivated strains of Porphyra haitanensis thalli(NB,PT and ST) were amplified,sequenced and analyzed.In addition,the phylogenic relationships of the sequences identified in this study with those of other Porphyra retrieved from GenBank were evaluated.The results are as follows:the sequences of the ITS and 5.8S rDNA were essentially identical among the three strains.The sequences of ITS1 were 331 bp to 334 bp,while those of the 5.8S rDNA were 158 bp and the sequences of ITS2 ranged from 673 bp to 681 bp.The sequences of the ITS had a high level of homology(up to 99.5%) with that of P.haitanensis(DQ662228) retrieved from GenBank,but were only approximately 50% homologous with those of other species of Porphyra.The results obtained when a phylogenetic tree was constructed coincided with the results of the homology analysis.These results suggest that the three cultivated strains of P.haitanensis evolved conservatively and that the ITS showed evolutionary consistency.However,the sequences of the ITS and 5.8S rDNA of different Porphyra species showed great variations.Therefore,the relationship of Porphyra interspecies phyletic evolution could be judged,which provides the proof for Porphyra identification study.However,proper classifications of the subspecies and the populations of Porphyra should be determined through the use of other molecular techniques to determine the genetic variability and rational phylogenetic relationships.展开更多
Two primers, designed according to the Arabidoposis thaliana Toc33 sequence, were used to amplify the full coding region of the Toc33 cDNAs in leaves of a chlorophyll-reduced (Cr) mutant Cr3529 and its wild type 3529,...Two primers, designed according to the Arabidoposis thaliana Toc33 sequence, were used to amplify the full coding region of the Toc33 cDNAs in leaves of a chlorophyll-reduced (Cr) mutant Cr3529 and its wild type 3529, Brassica napus, by RT-PCR technique. The RT-PCR results showed that the fragment homologous to Toc33 was expressed in Cr3529 as well as 3529 seedlings. PCR fragments were inserted into the pMD18-T vector and transferred into E. coli, then two cDNA clones, BnToc33-c and BnToc33, were obtained. Sequence analysis showed that the two sequences were 894 bp and the nucleotide and the deduced amino acid sequences were highly homologous to those of A. thaliana. There were three diverged nucleotides between the Cr3529 and the 3529 Toc33 cDNAs, i.e., GGT/AGT, TTG/TTT, AGG/AGT, all of which belonged to missense mutation. The amino acid replacement ((Leu/Phe) caused by TTG/TTT mutation located in the membrane anchor domain may result in chlorophyll-reduced character in Cr3529.展开更多
By screening the worker (Apis cerana cerana) heads cDNA library using a fragment of the mrjp3 gene of Apis cerana as probe, 120 positive clones were obtained. The clone containing A. cerana cerana MRJP3 (AccMRJP3)...By screening the worker (Apis cerana cerana) heads cDNA library using a fragment of the mrjp3 gene of Apis cerana as probe, 120 positive clones were obtained. The clone containing A. cerana cerana MRJP3 (AccMRJP3) cDNA was selected. Based on the sequencing of the inserts of the positive clone, a sequence of AccMRJP3 cDNA which is 1 887 bp long including a poly (A) tail was obtained. The AccMRJP3 cDNA encompassed an open-reading frame (ORF) with 1 779 bp encoding 593 amino acids. The un-translated regions (UTR) of the 5′ end and 3′end are 46 bp and 160 bp in length, respectively. Similar to AmMRJP3 and AdMRJP3, the putative AccMRJP3 also has a repetitive region. The comparison of the repetitive region of AccMRJP3, AmMRJP3 and AdMRJP3 shows some differences between them.展开更多
To confirm a hybrid swarm population of Pinus densiflora × P. sylvestris in Jilin, China, we used needles and seeds from P. densiflora, P. sylvestris, and P. densiflora × P. sylvestris collected from natural...To confirm a hybrid swarm population of Pinus densiflora × P. sylvestris in Jilin, China, we used needles and seeds from P. densiflora, P. sylvestris, and P. densiflora × P. sylvestris collected from natural stands or experimental stations to study whether shoot apex morphology of 4-year old seedlings can be correlated with the sequence of a chloroplast DNA simple sequence repeat marker (cpDNA SSRs). Total genomic DNA was extracted and subjected to sequence analysis of the pine cpDNA SSR marker Pt15169. Results show that morphological characters from 4-year old seedlings did not correlate with sequence variants of this marker. Marker haplotypes from all P. sylvestris trees had a CTAT element that was absent from all sampled P. densiflora trees. However, both haplotype classes involving this insertion/deletion element were found in a P. densiflora × P. sylvestris population and its seedling progeny. It was concluded that the P. densiflora × P. sylvestris accessions sampled from Jilin, China resulted from bi-directional crosses, as evidenced by both species’ cpDNA haplotypes within the hybrid swarm population.展开更多
Chitin is the most widespread amino polysaccharide in nature. Chitin synthase (CHS) plays an important role in chitin formation in the cuticle and the peritrophic membrane (PM) lining the midgut. Total RNA was iso...Chitin is the most widespread amino polysaccharide in nature. Chitin synthase (CHS) plays an important role in chitin formation in the cuticle and the peritrophic membrane (PM) lining the midgut. Total RNA was isolated from the cuticle of Mamestra brassicae (L.) fourth instar larva, cDNA sequence was cloned by RT-PCR and Rapid Amplification of cDNA Ends (RACE). cDNA 5 220 bp in length, contained an open reading frame of 4 704 bp coding for a polypeptide of 1 567 amino acid residues with a predicted molecular weight of 178.3 ku and its pI was 6.42. The deduced amino acid sequence from Mi brassicae (L.) shared the high level of identity with chitin synthase sequences from other insects, especially lepidopteran insects, cDNA sequence has been deposited with GenBank under accession No. GQ281761展开更多
Caryopteris incana is a perennial shrub distributed in the temperate zone of the East Asia. It is found in West Kyushu in Japan, where it is designated as an endangered species. Tsushima, Nagasaki, which experienced r...Caryopteris incana is a perennial shrub distributed in the temperate zone of the East Asia. It is found in West Kyushu in Japan, where it is designated as an endangered species. Tsushima, Nagasaki, which experienced repeated connection and fragmentation between the Korean Peninsula and Japan, is an island on the route along which C. incana moved to Japan from continental Asia. We conducted field work and confirmed the genetic structure of populations using DNA sequence analysis to construct a detailed distribution map and clarify the intraspecific phylogenetic relationships of C. incana in Tsushima Island. We confirmed 72 populations in Tsushima. Using the leaves of individuals cultivated from seeds collected from each natural population, we analyzed the chloroplast and nuclear DNA sequence variations. Among the populations, sequence variations were confirmed in six regions of chloroplast DNA, and six haplotypes, including base substitutions, were distinguished. Two haplotypes were mainly divided at the border of the northern part of the southern island in Tsushima. One population in the northwestern part of the north island showed a haplotype derived from the southern part. This finding revealed that the distribution of C. incana had been artificially influenced. Several haplotypes were confirmed by sequence variations in the northern populations, but only one haplotype in the southern populations, suggesting that C. incana on the north island had separated early from the south island in Tsushima.展开更多
Background: The multiple sequence alignment (MSA) algorithms are the traditional ways to compare and analyze DNA sequences. However, for large DNA sequences, these algorithms require a long time computationally. Objec...Background: The multiple sequence alignment (MSA) algorithms are the traditional ways to compare and analyze DNA sequences. However, for large DNA sequences, these algorithms require a long time computationally. Objective: Here we will propose a new numerical method to characterize and compare DNA sequences quickly. Method: Based on a new 2-dimensional (2D) graphical representation of DNA sequences, we can obtain an 8-dimensional vector using two basic concepts of probability, the mean and the variance. Results: We perform similarity/dissimilarity analyses among two real DNA data sets, the coding sequences of the first exon of beta-globin gene of 11 species and 31 mammalian mitochondrial genomes, respectively. Conclusion: Our results are in agreement with the existing analyses in our literatures. We also compare our approach with other methods and find that ours is more effective.展开更多
AIM: To study genetic difference of mitochondrial DNA (mtDNA)between two hepatocarcinoma cell lines (Hca-F and Hca-P)with diverse metastatic characteristics and the relationship between mtDNA changes in cancer cells a...AIM: To study genetic difference of mitochondrial DNA (mtDNA)between two hepatocarcinoma cell lines (Hca-F and Hca-P)with diverse metastatic characteristics and the relationship between mtDNA changes in cancer cells and their oncogenic phenotype.METHODS: Mitochondrial DNA D-loop, tRNAMet+Glu+Ile and ND3gene fragments from the hepatocarcinoma cell lines with 1100, 1126 and 534 bp in length respectively were analysed by PCR amplification and restriction fragment length polymorphism techniques. The D-loop 3' end sequence of the hepatocarcinoma cell lines was determined by sequencing.RESULTS: No amplification fragment length polymorphism and restriction fragment length polymorphism were observed in tRNAMet+Glu+Ile,ND3 and D-loop of mitochondrial DNA of the hepatocarcinoma cells. Sequence differences between Hca-F and Hca-P were found in mtDNA D-loop.CONCLUSION: Deletion mutations of mitochondrial DNA restriction fragment may not play a significant role in carcinogenesis. Genetic difference of mtDNA D-loop between Hca-F and Hca-P, which may reflect the environmental and genetic influences during tumor progression, could be linked to their tumorigenic phenotypes.展开更多
DNA methylation has been extensively investigated in recent years,not least because of its known relationship with various diseases.Progress in analytical methods can greatly increase the relevance of DNA methylation ...DNA methylation has been extensively investigated in recent years,not least because of its known relationship with various diseases.Progress in analytical methods can greatly increase the relevance of DNA methylation studies to both clinical medicine and scientific research.Microflu-idic chips are excellent carriers for molecular analysis,and their use can provide improvements from multiple aspects.On-chip molecular analysis has received extensive attention owing to its advantages of portability,high throughput,low cost,and high efficiency.In recent years,the use of novel microfluidic chips for DNA methylation analysis has been widely reported and has shown obvious superiority to conventional methods.In this review,wefirst focus on DNA methylation and its applications.Then,we discuss advanced microfluidic-based methods for DNA methylation analysis and describe the great progress that has been made in recent years.Finally,we summarize the advantages that microfluidic technology brings to DNA methylation analysis and describe several challenges and perspectives for on-chip DNA methylation analysis.This review should help researchers improve their understanding and make progress in developing microfluidic-based methods for DNA methylation analysis.展开更多
Wax gourd (Benincasa hispida Thumb. Cogn) is called white gourd, winter melon, Chinese preserving melon, Chinese squash, and don kwa. It has been cultivated in China for over 2 300 years. It probably
Melon downy mildew caused by Pseudoperonospora cubensis leads to significant losses in melon yields worldwide. Reverse-transcription Polymerase Chain Reaction (RT-PCR) was performed using cDNAs as templates from mel...Melon downy mildew caused by Pseudoperonospora cubensis leads to significant losses in melon yields worldwide. Reverse-transcription Polymerase Chain Reaction (RT-PCR) was performed using cDNAs as templates from melon-Huangdanzi induced with fungus Pseudoperonospora cubensis, and degenerate primers designed based on the conserved amino acid sequences of known plant disease-resistance genes. A polymorphic cDNA fragment which we named rap-19 was cloned and sequenced. The Open Reading Frame (ORF) of this product comprised of 510 base pairs which encodes DNA or RNA-binding protein with 170 amino acids. The putative amino acid sequence of mp-19 appeared highly homologous with those of NBS-type resistant-genes isolated from other plants. Southern blot indicated that the melon genome contained more than 3 copies of rap-19. The obvious expression differences detected by semi-quantitative RT- PCR could be observed between resistant-line Huangdanzi and susceptible-line Jiashi after Pseudoperonospora cubensis infection, which implied that mp-19 gene may be related to the resistance of downy mildew in melon.展开更多
BACKGROUND Genetic tests are increasingly performed for the management of unresectable pancreatic cancer.For genotyping aimed samples current guidelines recommend using core specimens,although based on moderate qualit...BACKGROUND Genetic tests are increasingly performed for the management of unresectable pancreatic cancer.For genotyping aimed samples current guidelines recommend using core specimens,although based on moderate quality evidence.However,in clinical practice among the endoscopic ultrasound(EUS) guided tissue acquisition methods,fine needle aspiration(FNA) is the most widely performed.AIM To assess the adequacy for next generation sequencing(NGS) of the DNA yielded from EUS-FNA pancreatic adenocarcinoma(PDAC) samples.METHODS Between November 2018 and December 2021,105 patients with PDAC confirmed by EUS-FNA were included in the study at our tertiary gastroenterology center.Either 22 gauge(G) or 19G FNA needles were used.One pass was dedicated to DNA extraction.DNA concentration and purity(A260/280,A260/230) were assessed by spectrophotometry.We assessed the differences in DNA parameters according to needle size and tumor characteristics(size,location) and the adequacy of the extracted DNA for NGS(defined as A260/280 ≥ 1.7,and DNA yield:≥ 10 ng for amplicon based NGS,≥ 50 ng for whole exome sequencing [WES],≥ 100 ng for whole genome sequencing [WGS]) by analysis of variance and ttest respectively.Moreover,we compared DNA purity parameters across the different DNA yield categories.RESULTS Our cohort included 49% male patients,aged 67.02 ± 8.38 years.The 22G needle was used in 71%of the cases.The DNA parameters across our samples varied as follows:DNA yield:1289 ng(inter quartile range:534.75-3101),A260/280 = 1.85(1.79-1.86),A260/230 = 2.2(1.72-2.36).DNA yield was > 10 ng in all samples and > 100 ng in 93% of them(one sample < 50 ng).There were no significant differences in the concentration and A260/280 between samples by needle size.Needle size was the only independent predictor of A260/230 which was higher in the 22G samples(P =0.038).NGS adequacy rate was 90% for 19G samples regardless of NGS type,and for 22G samples it reached 89% for WGS adequacy and 91% for WES and amplicon based NGS.Samples with DNA yield > 100 ng had significantly higher A260/280(1.89 ± 0.32 vs 1.34 ± 0.42,P = 0.013).Tumor characteristics were not corelated with the DNA parameters.CONCLUSION EUS-FNA PDAC samples yield DNA adequate for subsequent NGS.DNA amount was similar between 22G and 19G FNA needles.DNA purity parameters may vary indirectly with needle size.展开更多
基金The work was supported by the Hi-Tech Research and Development Program of China under contract Nos 2006AA09Z414 and 2007AA091903;the China Ocean Mineral Resources R & D Association under contract No. DYXM - 115 - 02 - 2 - 6;the National Natural Science Foundation of China under contract No. Z2004D02;the Natural Science Foundation of Shandong Province of China under contract No. Z2004D02;the Foundation for Young Excellent Scientists in Shandong Province of China under contract No. 2006BS02002;the Program for New Century Excellent Talents in University under contract No. NCET - 06 - 0578.
文摘Pseudoalteromonas sp. SM9913 is a phychrotmphic bacterium isolated from the deep-sea sediment. The genes encoding chaperones DnaJ and DnaK of P. sp. SM9913 were cloned by normal PCR and TAIL - PCR (GenBank accession Nos DQ640312, DQ504163 ). The chaperones DnaJ and DnaK from the strain SM9913 contain such conserved domains as those of many other bacteria, and show some cold-adapted characteristics in their structures when compared with those from psychro-, meso-and themophilic bacteria. It is indicated that chaperones DnaJ and DnaK of P. sp. SM9913 may be adapted to low temperature in deep-sea and function well in assisting folding, assembling and translocation of proteins at low temperature. This research lays a foundation for the further study on the cold-adapted mechanism of chaperones DnaJ and DnaK of cold-adapted microorganisms.
基金supported by funds from the National Natural Science Foundation of China(30800725 and30770278)the Central Public Research Institutes Basic Funds for Research and Development(Agro-Environmental Protection Institute,Ministry of Agriculture,China)
文摘The aim of this study was to isolate the full-length cDNA sequences of an unusually highly conserved olfactory receptor (orthologue to the Drosophila melanogaster DOR83b) from the antennae of Spodoptera exigua and S. litura by using reverse transcription-polymerase chain reaction (RT-PCR) and rapid amplification of cDNA ends (RACE) methods. Bioinformatics methods were used to further analyze the cDNA sequences and putative amino acid sequences. The two full-length cDNA sequences from olfactory receptor (OR) of male S. exigua and S. litura were named as SexiOR2 and SlitOR2, respectively. SexiOR2 and SlitOR2 consisted of nucleotide sequence of 1 906 and 2 483 bp, respectively, and both with deduced amino acid sequences of 473 residues. The sequence analysis indicated that the deduced amino acid sequences of the cDNA shared the high homologies with OR83b orthologue chemoreceptor sequences from previously reported moths, implying that the cDNA sequences were of OR83b orthologue chemoreceptor genes.
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.
基金Supported by the National Natural Science Foundation of China (No 40576074)the Key Laboratory of Experimental Marine Biology,Institute of Oceanology,Chinese Academy of Sciences (No KFN92007NO1)
文摘The sequences of the ITS(internal transcribed spacer) and 5.8S rDNA of three cultivated strains of Porphyra haitanensis thalli(NB,PT and ST) were amplified,sequenced and analyzed.In addition,the phylogenic relationships of the sequences identified in this study with those of other Porphyra retrieved from GenBank were evaluated.The results are as follows:the sequences of the ITS and 5.8S rDNA were essentially identical among the three strains.The sequences of ITS1 were 331 bp to 334 bp,while those of the 5.8S rDNA were 158 bp and the sequences of ITS2 ranged from 673 bp to 681 bp.The sequences of the ITS had a high level of homology(up to 99.5%) with that of P.haitanensis(DQ662228) retrieved from GenBank,but were only approximately 50% homologous with those of other species of Porphyra.The results obtained when a phylogenetic tree was constructed coincided with the results of the homology analysis.These results suggest that the three cultivated strains of P.haitanensis evolved conservatively and that the ITS showed evolutionary consistency.However,the sequences of the ITS and 5.8S rDNA of different Porphyra species showed great variations.Therefore,the relationship of Porphyra interspecies phyletic evolution could be judged,which provides the proof for Porphyra identification study.However,proper classifications of the subspecies and the populations of Porphyra should be determined through the use of other molecular techniques to determine the genetic variability and rational phylogenetic relationships.
基金This work was supported by the National Natural Science Foundation of China(30170500)the Training Foundation ofthe Academic Leader ofSichuan Province,P.R.China.
文摘Two primers, designed according to the Arabidoposis thaliana Toc33 sequence, were used to amplify the full coding region of the Toc33 cDNAs in leaves of a chlorophyll-reduced (Cr) mutant Cr3529 and its wild type 3529, Brassica napus, by RT-PCR technique. The RT-PCR results showed that the fragment homologous to Toc33 was expressed in Cr3529 as well as 3529 seedlings. PCR fragments were inserted into the pMD18-T vector and transferred into E. coli, then two cDNA clones, BnToc33-c and BnToc33, were obtained. Sequence analysis showed that the two sequences were 894 bp and the nucleotide and the deduced amino acid sequences were highly homologous to those of A. thaliana. There were three diverged nucleotides between the Cr3529 and the 3529 Toc33 cDNAs, i.e., GGT/AGT, TTG/TTT, AGG/AGT, all of which belonged to missense mutation. The amino acid replacement ((Leu/Phe) caused by TTG/TTT mutation located in the membrane anchor domain may result in chlorophyll-reduced character in Cr3529.
基金the National Natural Science Foundation of China(30200206) Zhejiang Provincial Natural Science Foundation of China(302113).
文摘By screening the worker (Apis cerana cerana) heads cDNA library using a fragment of the mrjp3 gene of Apis cerana as probe, 120 positive clones were obtained. The clone containing A. cerana cerana MRJP3 (AccMRJP3) cDNA was selected. Based on the sequencing of the inserts of the positive clone, a sequence of AccMRJP3 cDNA which is 1 887 bp long including a poly (A) tail was obtained. The AccMRJP3 cDNA encompassed an open-reading frame (ORF) with 1 779 bp encoding 593 amino acids. The un-translated regions (UTR) of the 5′ end and 3′end are 46 bp and 160 bp in length, respectively. Similar to AmMRJP3 and AdMRJP3, the putative AccMRJP3 also has a repetitive region. The comparison of the repetitive region of AccMRJP3, AmMRJP3 and AdMRJP3 shows some differences between them.
基金supported by a grant from the Next-Generation BioGreen 21 Program, Rural Development Administration, Republic of Korea (PJ009052)
文摘To confirm a hybrid swarm population of Pinus densiflora × P. sylvestris in Jilin, China, we used needles and seeds from P. densiflora, P. sylvestris, and P. densiflora × P. sylvestris collected from natural stands or experimental stations to study whether shoot apex morphology of 4-year old seedlings can be correlated with the sequence of a chloroplast DNA simple sequence repeat marker (cpDNA SSRs). Total genomic DNA was extracted and subjected to sequence analysis of the pine cpDNA SSR marker Pt15169. Results show that morphological characters from 4-year old seedlings did not correlate with sequence variants of this marker. Marker haplotypes from all P. sylvestris trees had a CTAT element that was absent from all sampled P. densiflora trees. However, both haplotype classes involving this insertion/deletion element were found in a P. densiflora × P. sylvestris population and its seedling progeny. It was concluded that the P. densiflora × P. sylvestris accessions sampled from Jilin, China resulted from bi-directional crosses, as evidenced by both species’ cpDNA haplotypes within the hybrid swarm population.
基金Supported by Natural Science Foundation of Heilongjiang Province (C2007-7)Scientific and Technical Innovation Fund of Harbin (RC2006QN002027)Northeast Agricultural University Research Fund (2005)
文摘Chitin is the most widespread amino polysaccharide in nature. Chitin synthase (CHS) plays an important role in chitin formation in the cuticle and the peritrophic membrane (PM) lining the midgut. Total RNA was isolated from the cuticle of Mamestra brassicae (L.) fourth instar larva, cDNA sequence was cloned by RT-PCR and Rapid Amplification of cDNA Ends (RACE). cDNA 5 220 bp in length, contained an open reading frame of 4 704 bp coding for a polypeptide of 1 567 amino acid residues with a predicted molecular weight of 178.3 ku and its pI was 6.42. The deduced amino acid sequence from Mi brassicae (L.) shared the high level of identity with chitin synthase sequences from other insects, especially lepidopteran insects, cDNA sequence has been deposited with GenBank under accession No. GQ281761
文摘Caryopteris incana is a perennial shrub distributed in the temperate zone of the East Asia. It is found in West Kyushu in Japan, where it is designated as an endangered species. Tsushima, Nagasaki, which experienced repeated connection and fragmentation between the Korean Peninsula and Japan, is an island on the route along which C. incana moved to Japan from continental Asia. We conducted field work and confirmed the genetic structure of populations using DNA sequence analysis to construct a detailed distribution map and clarify the intraspecific phylogenetic relationships of C. incana in Tsushima Island. We confirmed 72 populations in Tsushima. Using the leaves of individuals cultivated from seeds collected from each natural population, we analyzed the chloroplast and nuclear DNA sequence variations. Among the populations, sequence variations were confirmed in six regions of chloroplast DNA, and six haplotypes, including base substitutions, were distinguished. Two haplotypes were mainly divided at the border of the northern part of the southern island in Tsushima. One population in the northwestern part of the north island showed a haplotype derived from the southern part. This finding revealed that the distribution of C. incana had been artificially influenced. Several haplotypes were confirmed by sequence variations in the northern populations, but only one haplotype in the southern populations, suggesting that C. incana on the north island had separated early from the south island in Tsushima.
文摘Background: The multiple sequence alignment (MSA) algorithms are the traditional ways to compare and analyze DNA sequences. However, for large DNA sequences, these algorithms require a long time computationally. Objective: Here we will propose a new numerical method to characterize and compare DNA sequences quickly. Method: Based on a new 2-dimensional (2D) graphical representation of DNA sequences, we can obtain an 8-dimensional vector using two basic concepts of probability, the mean and the variance. Results: We perform similarity/dissimilarity analyses among two real DNA data sets, the coding sequences of the first exon of beta-globin gene of 11 species and 31 mammalian mitochondrial genomes, respectively. Conclusion: Our results are in agreement with the existing analyses in our literatures. We also compare our approach with other methods and find that ours is more effective.
基金Supported by the National Natural Science Foundation of China, No. 39900173
文摘AIM: To study genetic difference of mitochondrial DNA (mtDNA)between two hepatocarcinoma cell lines (Hca-F and Hca-P)with diverse metastatic characteristics and the relationship between mtDNA changes in cancer cells and their oncogenic phenotype.METHODS: Mitochondrial DNA D-loop, tRNAMet+Glu+Ile and ND3gene fragments from the hepatocarcinoma cell lines with 1100, 1126 and 534 bp in length respectively were analysed by PCR amplification and restriction fragment length polymorphism techniques. The D-loop 3' end sequence of the hepatocarcinoma cell lines was determined by sequencing.RESULTS: No amplification fragment length polymorphism and restriction fragment length polymorphism were observed in tRNAMet+Glu+Ile,ND3 and D-loop of mitochondrial DNA of the hepatocarcinoma cells. Sequence differences between Hca-F and Hca-P were found in mtDNA D-loop.CONCLUSION: Deletion mutations of mitochondrial DNA restriction fragment may not play a significant role in carcinogenesis. Genetic difference of mtDNA D-loop between Hca-F and Hca-P, which may reflect the environmental and genetic influences during tumor progression, could be linked to their tumorigenic phenotypes.
基金support from the National Key R&D Program of China(Grant No.2018YFE0118700)the National Natural Science Foundation of China(NSFC Grant No.62174119)+1 种基金the 111 Project(Grant No.B07014)the Foundation for Talent Scientists of Nanchang Institute for Microtechnology of Tianjin University.
文摘DNA methylation has been extensively investigated in recent years,not least because of its known relationship with various diseases.Progress in analytical methods can greatly increase the relevance of DNA methylation studies to both clinical medicine and scientific research.Microflu-idic chips are excellent carriers for molecular analysis,and their use can provide improvements from multiple aspects.On-chip molecular analysis has received extensive attention owing to its advantages of portability,high throughput,low cost,and high efficiency.In recent years,the use of novel microfluidic chips for DNA methylation analysis has been widely reported and has shown obvious superiority to conventional methods.In this review,wefirst focus on DNA methylation and its applications.Then,we discuss advanced microfluidic-based methods for DNA methylation analysis and describe the great progress that has been made in recent years.Finally,we summarize the advantages that microfluidic technology brings to DNA methylation analysis and describe several challenges and perspectives for on-chip DNA methylation analysis.This review should help researchers improve their understanding and make progress in developing microfluidic-based methods for DNA methylation analysis.
文摘Wax gourd (Benincasa hispida Thumb. Cogn) is called white gourd, winter melon, Chinese preserving melon, Chinese squash, and don kwa. It has been cultivated in China for over 2 300 years. It probably
基金This study was supported by the Natural Science Foun-dation of Xinjiang Uygur Autonomous Region, China(200121105)Science and Research Plan Emphasis Project of College in Xinjiang Uygur Autonomous Region, China (XJEDU2004I09).
文摘Melon downy mildew caused by Pseudoperonospora cubensis leads to significant losses in melon yields worldwide. Reverse-transcription Polymerase Chain Reaction (RT-PCR) was performed using cDNAs as templates from melon-Huangdanzi induced with fungus Pseudoperonospora cubensis, and degenerate primers designed based on the conserved amino acid sequences of known plant disease-resistance genes. A polymorphic cDNA fragment which we named rap-19 was cloned and sequenced. The Open Reading Frame (ORF) of this product comprised of 510 base pairs which encodes DNA or RNA-binding protein with 170 amino acids. The putative amino acid sequence of mp-19 appeared highly homologous with those of NBS-type resistant-genes isolated from other plants. Southern blot indicated that the melon genome contained more than 3 copies of rap-19. The obvious expression differences detected by semi-quantitative RT- PCR could be observed between resistant-line Huangdanzi and susceptible-line Jiashi after Pseudoperonospora cubensis infection, which implied that mp-19 gene may be related to the resistance of downy mildew in melon.
基金The Executive Agency for Higher Education,Research,Development and Innovation Funding-research,No.PN-Ⅲ-P1-1.2-PCCDI-2017-0797 (PANCNGS)
文摘BACKGROUND Genetic tests are increasingly performed for the management of unresectable pancreatic cancer.For genotyping aimed samples current guidelines recommend using core specimens,although based on moderate quality evidence.However,in clinical practice among the endoscopic ultrasound(EUS) guided tissue acquisition methods,fine needle aspiration(FNA) is the most widely performed.AIM To assess the adequacy for next generation sequencing(NGS) of the DNA yielded from EUS-FNA pancreatic adenocarcinoma(PDAC) samples.METHODS Between November 2018 and December 2021,105 patients with PDAC confirmed by EUS-FNA were included in the study at our tertiary gastroenterology center.Either 22 gauge(G) or 19G FNA needles were used.One pass was dedicated to DNA extraction.DNA concentration and purity(A260/280,A260/230) were assessed by spectrophotometry.We assessed the differences in DNA parameters according to needle size and tumor characteristics(size,location) and the adequacy of the extracted DNA for NGS(defined as A260/280 ≥ 1.7,and DNA yield:≥ 10 ng for amplicon based NGS,≥ 50 ng for whole exome sequencing [WES],≥ 100 ng for whole genome sequencing [WGS]) by analysis of variance and ttest respectively.Moreover,we compared DNA purity parameters across the different DNA yield categories.RESULTS Our cohort included 49% male patients,aged 67.02 ± 8.38 years.The 22G needle was used in 71%of the cases.The DNA parameters across our samples varied as follows:DNA yield:1289 ng(inter quartile range:534.75-3101),A260/280 = 1.85(1.79-1.86),A260/230 = 2.2(1.72-2.36).DNA yield was > 10 ng in all samples and > 100 ng in 93% of them(one sample < 50 ng).There were no significant differences in the concentration and A260/280 between samples by needle size.Needle size was the only independent predictor of A260/230 which was higher in the 22G samples(P =0.038).NGS adequacy rate was 90% for 19G samples regardless of NGS type,and for 22G samples it reached 89% for WGS adequacy and 91% for WES and amplicon based NGS.Samples with DNA yield > 100 ng had significantly higher A260/280(1.89 ± 0.32 vs 1.34 ± 0.42,P = 0.013).Tumor characteristics were not corelated with the DNA parameters.CONCLUSION EUS-FNA PDAC samples yield DNA adequate for subsequent NGS.DNA amount was similar between 22G and 19G FNA needles.DNA purity parameters may vary indirectly with needle size.