Sequence Analysis of Mitochondrial DNA D-loop Region in Xinjiang Goose

[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）.

Cloning, sequencing and analyzing of the heavy chain V region genes of human polyreactive antibodies

The heavy chain variable region genes of 5 human polyreactive mAbs generated in our laboratory have been cloned and sequenced using polymerase chain reaction (PCR) technique. We found that 2 and 3 mAbs utilized genes of the VHIV and VHIII families, respectively. The former 2 VH segments were in germline configuration. A common VH segment, with the best similarity of 90.1 % to the published VHIII germline genes, was utilized by 2 different rearranged genes encoding the V regions of other 3 mAbs. This strongly suggests that the common VH segment is a unmutated copy of an unidentified germline VHIII gene. All these polyreactive mAbs displayed a large NDN region (VH-D-JH junction). The entire H chain V regions of these polyreactive mAbs are unusually basic. The analysis of the charge properties of these mAbs as well as those of other poly- and mono- reactive mAbs from literatures prompts us to propose that the charged amino acids with a particular distribution along the H chain V region,especially the binding sites (CDRs), may be an important structural feature involved in antibody polyreactivity.

Sequence Difference of Mitochondrial DNA Control Region and Genetic Differentiation of Cervinae in China

[Objective] The study aimed at analyzing Genetic Differentiation of Cervinae. [Method] The complete mitochondrial DNA control region of five species of Cervinae was determined by direct DNA sequencing. Homologous sequences of others Cervinae were gained from GenBank, and then analyzed through biology software. [Result] The lengths of their control region were 921-1 072 bp, the nucleotide content of T, A, C, and G was 32.1%, 30.2%, 22.7% and 15.0% separately. Genetic distance of each species ranged from 0.062-0.106, which belonged to level of inter-genus. The molecular phylogenetic tree indicated that red deer, skia deer and White-lipped Deer had close relationship, and they constituted one branch with sambar deer. Eld＇s deer and elk deer constituted the second branch, and hog deer constituted the third branch alone. [Conclusion] Hog deer and fallow deer should be incorporated in Cervus, divergence time is about 1.55-2.60 million years among Cervinae in China.

Large-scale Migration of Fluids toward Foreland Basins during Collisional Orogeny: Evidence from Triassic Anhydrock Sequences and Regional Alteration in the Middle-Lower Yangtze Area

The middle-lower Yangtze area underwent a series of complex tectonic evolution, such as Hercynian extensional rifting, Indosinian foreland basining, and Yanshanian transpression-transtension, resulting in a large distinctive Cu-Fe-Au metallogenic belt. In the tectonic evolution, large-scale migration and convergence of fluids toward foreland basins induced during the collisional orogeny of the Yangtze and North China continental blocks were of vital importance for the formation of the metallogenic belt. Through geological surveys of the middle-lower Yangtze area, three lines of evidence of large-scale fluid migration are proposed: (1) The extensive dolomitic and silicic alteration penetrating Cambrian-Triassic strata generally occurs in a region sandwiched between the metallogenic belt along the Yangtze River and the Dabie orogenic belt, and in the alteration domain alternately strong and weak alteration zones extend in a NW direction and are controlled by the fault system of the Dabie orogenic belt; it might record the locus of the activities of long-distance migrating fluids. (2) The textures and structures of very thick Middle-Lower Triassic anhydrock sequences in restricted basins along the river reveal the important contribution of the convergence of regional hot brine in restricted basins and the chemical deposition or their formation. (3) Early-Middle Triassic syndepositional iron carbonate sequences and Fe-Cu-Pb-Zn massive sulfide deposits alternate with anhydrock sequences or are separated from the latter, but all of them occur in the same stratigraphic horizon and are intimately associated with each other, being the product of syndeposition of high-salinity hot brine. According to the geological surveys, combined with previous data, the authors propose a conceptual model of fluid migration-convergence and mineralization during the Dabie collisional orogeny.

Development of Sequence Characterized Amplified Region (SCAR) Primers for the Detection of Resistance to Sporisorium reiliana in Maize

Head smut of maize （Zea mays L.）, which was caused by Sporisorium reiliana, occurred in most of the maize growing areas of the world. The purpose of this study was to develop SCAR markers for map-based cloning of resistance genes and MAS. Two sets of BC3 progenies, one （BC3Q） derived from the cross Qi319 （resistance）×Huangzao 4 （susceptible）, the other （BC3M） from Mol7 （resistance）× Huangzao 4 （susceptible）, were generated. Huangzao 4 was the recurrent parent in both progenies. A combination of BSA （bulked segregant analysis） with AFLP （amplified fragment length polymorphism） method was applied to map the genes involving the resistance to S. reiliana, and corresponding resistant and susceptible bulks and their parental lines were used for screening polymorphic AFLP primer pairs. One fragment of PI3M61-152 was converted into SCAR （sequence charactered amplified fragment） marker S130. The marker was mapped at chromosome bin 2.09, the interval of a major QTL region previously reported to contribute to S. reiliana resistance. Furthermore, S130 was highly and facilitate map-based cloni associated with resistance to S. reiliana, and could be useful for marker-assisted selection ng of resistance genes.

The Hyperspace of the Regions Below of Continuous Maps from the Converging Sequence

Let S = {1,1/2,1/2^2,…,1/∞ = 0} and I = [0, 1] be the unit interval. We use ↓USC（S） and ↓C（S） to denote the families of the regions below of all upper semi-continuous maps and of the regions below of all continuous maps from S to I and ↓C0（S） = {↓f∈↓C（S） ： f（0） = 0}. ↓USC（S） endowed with the Vietoris topology is a topological space. A pair of topological spaces （X, Y） means that X is a topological space and Y is its subspace. Two pairs of topological spaces （X, Y） and （A, B） are called pair-homeomorphic （≈） if there exists a homeomorphism h ： X→A from X onto A such that h（Y） = B. It is proved that, （↓USC（S）,↓C0（S）） ≈（Q, s） and （↓USC（S）,↓C（S）／ ↓C0（S））≈（Q, c0）, where Q = [-1,1]^ω is the Hilbert cube and s = （-1,1）^ω,c0= {（xn）∈Q ： limn→∞= 0}. But we do not know what （↓USC（S）,↓C（S））is.

Sequence analysis of the ITS region and 5.8S rDNA of Porphyra haitanensis

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.

MicroRNA and mRNA profiling of cerebral cortex in a transgenic mouse model of Alzheimer’s disease by RNA sequencing

In a previous study,we found that long non-coding genes in Alzheimer’s disease(AD)are a result of endogenous gene disorders caused by the recruitment of microRNA(miRNA)and mRNA,and that miR-200a-3p and other representative miRNAs can mediate cognitive impairment and thus serve as new biomarkers for AD.In this study,we investigated the abnormal expression of miRNA and mRNA and the pathogenesis of AD at the epigenetic level.To this aim,we performed RNA sequencing and an integrative analysis of the cerebral cortex of the widely used amyloid precursor protein and presenilin-1 double transgenic mouse model of AD.Overall,129 mRNAs and 68 miRNAs were aberrantly expressed.Among these,eight down-regulated miRNAs and seven up-regulated miRNAs appeared as promising noninvasive biomarkers and therapeutic targets.The main enriched signaling pathways involved mitogen-activated kinase protein,phosphatidylinositol 3-kinase-protein kinase B,mechanistic target of rapamycin kinase,forkhead box O,and autophagy.An miRNA-mRNA network between dysregulated miRNAs and corresponding target genes connected with AD progression was also constructed.These miRNAs and mRNAs are potential biomarkers and therapeutic targets for new treatment strategies,early diagnosis,and prevention of AD.The present results provide a novel perspective on the role of miRNAs and mRNAs in AD.This study was approved by the Experimental Animal Care and Use Committee of Institute of Medicinal Biotechnology of Beijing,China(approval No.IMB-201909-D6)on September 6,2019.

Rapid detection of self-biting disease of mink by specific sequence-characterized amplified regions

Self-biting disease occurred in most farmed fur animals in the world. The mechanism and rapid detection method of this disease has not been reported. We applied bulked sergeant analysis （BSA） in combination with RAPD method to analyze a molecular genetic marker linked with self-biting trait in mink group. The molecular marker was converted into sequence-characterized amplified regions （SCAR） marker for rapid detection of this disease. A single RAPD marker A8 amplified a specific band of 263bp in self-biting minks, which was designated as SRA8-250, and non-specific band of 315bp in both self-biting and healthy minks. The sequences of the bands exhibited 75% and 88% similarity to Canis familiarizes major histocompatibility complex （MHC） class II region and Macaca mulatta MHC class I region, respectively. A SCAR marker SCAR-A8 was designed for the specific fragment SRA8-250 and validated in 30 self-biting minks and 30 healthy minks. Positive amplification of SCAR-A8 was detected in 24 self-biting minks and 12 healthy minks. χ2 test showed significant difference （p〈0.01） in the detection rate between the two groups. This indicated that SRA8-250 can be used as a positive marker to detect self-biting disease in minks. Furthermore, the finding that self-biting disease links with MHC genes has significant implications for the mechanism of the disease.

Panax ginseng-specific sequence characterized amplified region (SCAR) marker for testing medicinal products

To screen genetic polymorphisms of Panax ginseng, as well as those of Panax quinquefolium and Panax notoginseng, analysis of random amplified polymorphic DNA （RAPD） was performed using 120 random primers. Of the successful amplicons obtained, the Panax ginseng-specific RAPD marker C-12 was cloned into a TA vector and sequenced （Genl3ank access number KU553472）. Based on the sequence analysis results, a pair of primers specific to C-12 was designed. Finally, a SCAR marker-based identification system for Panax ginseng was developed after optimization of the reaction conditions. Using this method, two positive bands were stably observed at 300 bp and 130 bp in 33 batches of Panax ginseng samples tested, while negative results were obtained for another 101 batches of samples, including Panax quinquefolium, Panax notoginseng, adulterants, and other medicinal herbs. Thus, we successfully developed a PCR-based method for rapid and effective identification of Panax ginseng, which can be effectively used for the protection and utilization of germplasm resources and identification of the origins of Panax ginseng samples.

From Basin Black Shales to Platform Carbonate Rocks:A Study on Sequence Stratigraphy for the Lower Cambrian of the Upper-Yangtze Region in South China

In the Upper-Yangtze region, especially in Guizhou Province and its adjacent areas, the Lower Cambrian is well developed and is marked by a succession from black shales of the basin facies to carbonate rocks of the platform facies. The drowning event of the platform occurring at the turn from Sinian to Cambrian resulted in a set of black shales, i.e. the Niutitang Formation, which makes up the bottom part of the Lower Cambrian. With the shoaling of the sedimentary environment, a set of carbonate rocks, i.e. the Qingxudong Formation, was formed in the top part of the Lower Cambrian. Thus, the Lower Cambrian in the study area makes up one second-order sequence that can be further subdivided into five third-order sequences, and forms a regularly cyclic succession of transgression-regression. There is a regularly vertical stacking pattern for the third-order sequences in the second-order sequence. From bottom to top, the succession of the ＂CS （condensed section） ＋HST （high-stand system tract）＂ of the third-order sequences is changed into the succession of the ＂TST （transgressive system tract）＋CS＋HST＂. Correspondingly, the drowning-type sequence boundary is changed into the exposure-type one. Therefore, both the second-order and the third-order sequences have similar sedimentary-facies architectures. A concomitant with these temporal changes, the Lower Cambrian with a thickness of 1000 m that contains five third-order sequences is changed into a condensed succession that cannot identify third-order sequences toward the southeast with the deepening of the sedimentary environment. According to the elementary features of the third-order sequences, i.e. the regularity o sedimentary-facies successions in space and the synchronism of sedimentary-environment changes in time, the detailed division of the third-order sequences at main logged sections in different paleogeographical background becomes the basis to establish the sequence-stratigraphic framework that can demonstrate two types of facies-changing surface and two types of diachronism in the stratigraphic records. This sequence-stratigraphic framework shows a growing process of the carbonate platform from the base of the condensed succession formed by black shales of basin facies. Resulting from the rapid transgression at the turn from Sinian to Cambrian the ecological space became open, which formed the antecedent condition of paleogeographical setting for ＂the Cambrian Biological Explosion＂. Ultimately, the genetic relationship between the depositional events and the biological-diversity events is very complex and there remain lots of problems that need further research in the future.

The Late Triassic Sequence-Stratigraphic Framework of the Upper Yangtze Region,South China

In the transitional period between the Middle and the Late Triassic, the Indochina orogeny caused two tectonic events in South China：（1） the formation and uplift of the Qinling-Dabie orogenic belt along the northern margin of the South China Plate, due to its collision with the North China Plate; and 2） the development of a 1300-km-wide intra-continental orogen in the southeastern part of the South China Plate, which led to a northwestward movement of the foreland thrust-fold zone. These tectonic events resulted in the ending of the Yangtze Platform, and were a stable paleogeographic factor from the Eidacaran to the end of the Middle Triassic. This platform was characterized by the widespread development of shallow-water carbonates. After the end of the Yangtze Platform, the upper Yangtze foreland basin（or Sichuan foreland basin） was formed during the Late Triassic and became a accumulation site of fluvial deposits that are composed of related strata of the Xujiahe Formation. In western Sichuan Province, the Xujiahe Formation overlies the Maantang Formation shallow-water carbonate rocks of the Xiaotangzi Formation siliciclastic rocks（from shelf shales to littoral facies）. The sequence-stratigraphic framework of the Upper Triassic in the upper Yangtze foreland basin indicates a particular alluvial architecture, characterized by sequences composed of（1） successions of low-energy fluvial deposits of high-accommodation phases, including coal seams, and（2） high-energy fluvial deposits of low-accommodation phases, including amalgamated river-channel sandstones. The spatial distribution of these fluvial deposits belonging to the Xujiahe Formation and its relative strata is characterized by gradual thinning-out, overlapping, and pinching-out toward both the east and south. This sedimentary record therefore expresses a particular sequence-stratigraphic succession of fluvial deposits within the filling succession of the foreland basin. The sequence-stratigraphic framework for the Upper Triassic in the Upper Yangtze region provides a record of the end of the Yangtze Platform and the formation of the upper Yangtze foreland basin.

Research on Outcrop Sequence Stratigraphy of Permian in Middle-Lower Yangtze Region

According to the latest International Chronostratigraphic Scheme (ICS, 2000), the Permian in the Middle Lower Yangtze region of South China can be divided into three series and nine stages relevant to the traditional six stages of South China. From Assellian to Changxingian of Permian, 44 Ma in age range, the strata are composed of 14 third order sequences, each of which is 3.14 Ma in average age range. There is one third order sequence of Zisongian, equivalent to middle and upper Chuanshan Formation or equal to Asselian and two thirds of Sakmarian. There are two third order sequences, corresponding to Liang shan Formation or Zhenjiang Formation and upper Chuanshan Formation, which are assigned to Longlingian, coinciding with Artinskian and one third of Sakmarian. In addition, three third order sequences, equal to Qixia Formation, are attributed to Chihsian, corresponding to Kubergandian and one third of Roadian. Four third order sequences, comprising Gufeng, Maokou, Yanqiao, Yinping and Wuxue formations, are assigned to Maokouan, equivalent to two thirds of Roadian, Wordian and Capitanian. Two third order sequences, equal to Longtan Formation or Wujiaping Formation, are included in Wuchiapingian. Other two third order sequences, corresponding to Changxing Formation or Dalong Formation, are assigned to Changhsingian. In brief, these above third order sequences can be incorporated into 4 sequences sets.

Comparative Analysis of Mitochondrial Control Region Sequence from Three Flatfish Species(Pleuronectidae)

The 5’-end of the mitochondrial control region sequences of three flatfishes (Pleuronectiformes: Pleuronectidae) were amplified and sequenced. These sequences were compared with those of other three Pleuronectids species retrieved from GenBank. A phylogenetic tree was constructed based on the partial control region sequences. The results of phyloge- netic analysis are consistent with those of conventional systematics. Compared to previous studies, the structure of the 5’-end of mitochondrial control region was analyzed. The terminal associated sequence motif and its complementary motif were i- dentified at the 5’-end of the sequences. A conserved sequence block, named as CM5’d, was identified in the 5’-end of con- trol region sequences in all Pleuronectids. Another central conserved sequence block, named as CSB-F, was detected in the central conserved blocks.

Primary Analysis on mtDNA D-loop Hypervariable Region in Eutamias sibiricus

This study analyzed the mitochondrial DNA D-loop hypervariable region 601 bp sequence in 12 Eutamias sibiricus from Heilongjiang area. The result showed that the average contents of A, T, G and C were 33.2%, 30.5%, 11.8% and 24.5% respectively, the A＋T content （63.7%） was obviously higher than the G＋C content （36.3%）. Thirty-six, mutation （approximately 6.0%） sites were found and 9 haplotypes were defined. The mutations types, including transition, transversion and deletion were all found in the detected mtDNA D-loop regions, most of which was transition. The average nucleotide mutational ratio was 1.22%. The nucleotide mutation sites affected the restriction site appearance or disappearance of the restriction site. The research on mtDNA D-loop is focused on the domestic animals and there is no report on Eutamias sibiricus, This study analyzed the mitochondrial DNA D-loop hypervariable in Eutamias sibiricus so as to provide some useful informations for related research in the future.

Identification of the interactive region by the homology of the sequence spectrum

The base sequence in genome was governed by some fundamental principles such as reverse-complement symmetry, multiple fractality and so on, and the analytical method of the genome structure, the “Sequence Spectrum Method (SSM)”, based on the structural features of genomic DNA faithfully visualized these principles. This paper reported that the sequence spectrum in SSM closely reflected the biological phenomena of protein and DNA, and SSM could identify the interactive region of protein-protein and DNA-protein uniformly. In order to investigate the effectiveness of SSM we analyzed the several protein-protein and DNA-protein interaction published primarily in the genome of Saccharomyces cerevisiae. The method proposed here was based on the homology of sequence spectrum, and it advantageously and surprisingly used only base sequence of genome and did not require any other information, even information about the amino-acid sequence of protein. Eventually it was concluded that the fundamental principles in genome governed not only the static base sequence but also the dynamic function of protein and DNA.

Complete Sequence Determination and Origin of Mitochondrial DNA Control Region in Dehua Black Chicken

Primers were designed according to the genomic DNA of red jungle fowl, and the complete sequence of mtDNA D-loop region in Dehua black chicken was obtained. Combined with the D-loop region complete sequence of red jungle fowl in GenBank, the maternal origin of Dehua black chicken was explored. The results showed that the complete sequence of mtDNA D-loop region in Dehua black chicken was 1 231 bp, and the average contents of T, C, A and G were 33.5%, 26.5%, 26.7% and 13.3%, respectively. Phylogenetic analysis showed that Dehua black chicken clustered with red jungle fowl （ C, allus gaUus spadiceus） , suggesting Dehua black chicken probably originated from G. g. spadiceu~ in Yunnan Province or nearby areas.

Cloning and Sequence Analysis of Light Variable Region Gene of Anti-human Retinoblastoma Monoclonal Antibody

Purpose: To clone the variable region gene of light chain of monoclonal antibody against human retinoblastoma and to analyze the characterization of its nucleotide sequence as well as amino acid sequence.Methods: Total RNA was extracted from 3C6 hybridoma cells secreting specific monoclonal antibody(McAb)against human retinoblastoma(RB), then transcripted reversely into cDNA with olig-dT primers.The variable region of the light chain (VL) gene fragments was amplified using polymeerase chain reaction(PCR) and further cloned into pGEM(R) -T Easy vector. Then, 3C6 VL cDNA was sequenced by Sanger's method.Homologous analysis was done by NCBI BLAST.Results: The complete nucleotide sequence of 3C6 VL cDNA consisted of 321 bp encoding 107 amino acid residues, containing four workframe regions(FRs)and three complementarity-determining regions (CDRs) as well as the typical structure of two cys residues. The sequence is most homological to a member of the Vk9 gene family, and its chain utilizes the Jkl gene segment.Conclusion: The light chain variable region gene of the McAb against human RB was amplified successfully , which belongs to the Vk9 gene family and utilizes Vk-Jk1 gene rearrangement. This study lays a good basis for constructing a recombinant antibody and for making a new targeted therapeutic agents against retinoblastoma.

4种鼢鼠线粒体DNA D-loop区全序列分析

采用PCR测序技术对采自甘肃境内的4种鼢鼠22个个体的线粒体DAN D-loop区全序列进行了测定。结果表明:鼢鼠线粒体DNA D-loop序列长度为893、894、895、896或899 bp,核苷酸位点突变类型有5种,即转换、颠换、插入、缺失及转换与颠换共存。碱基转换以T<=>C形式为主。其中T、C、A、G 4种核苷酸的平均比例分别为34.1%、24.3%、30.1%、11.5%,A+T含量(64.2%)明显高于G+C含量(35.8%)。这4种鼢鼠22条线粒体DNA D-loop区发现20种单倍型,单倍型比例为81.82%,说明中国鼢鼠mtDNA遗传多态性很丰富。从系统发育树分析,4种鼢鼠明显聚为两类,推测鼢鼠可能有两个起源。