Sinorhizobium meliloti lsrB is involved in alfalfa root nodule development and nitrogen-fixing bacteroid differentiation

Rhizobia interact with host legumes to induce the formation of nitrogen-fixing nodules, which is very important in agriculture and ecology. The development of nitrogen-fixing nodules is stringently regulated by host plants and rhizobial symbionts. In our previous work, a new Sinorhizobium meliloti LysR regulator gene (lsrB) was identified to be essential for alfalfa nodulation. However, how this gene is involved in alfalfa nodulation was not yet understood. Here, we found that this gene was associated with prevention of premature nodule senescence and abortive bacteroid formation. Heterogeneous deficient alfalfa root nodules were induced by the in-frame deletion mutant of lsrB (lsrB1-2), which was similar to the plasmid-insertion mutant, lsrB1. Irregular senescence zones earlier appeared in these nodules where bacteroid differentiation was blocked at different stages from microscopy observations. Interestingly, oxidative bursts were observed in these nodules by DAB staining. The decreased expression of lipopolysaccharide core genes (lpsCDE) was correspondingly determined in these nodules. S. meliloti lipopolysaccharide is required for suppression of oxidative bursts or host cell defense. These findings demonstrate that the S. meliloti lsrB gene is involved in alfalfa root nodule development and bacteroid differentiation by suppressing oxidative bursts or defense responses in host cells.     

Cytogenetic Mapping of Disease Resistance Genes and Analysis of Their Distribution Features on Chromosomes in Maize

0　IntroductionMaizeisamongthemostintensivelystudiedspeciesingeneticsandoneofagronomicallythemostimportantplants.Therearemanydis easemicrobesandpeststoattackmaize,whichre sultsinlowproductionandbadquality .Withthedevelopmentofverydensegeneticmapconstruc tion ,avarietyoftheimportantdiseaseresistancegenesofmaizeincludingHelminthosporiumtur ciumPassresistancegenesHt1,Htn1andHt2 ,HelminthosporiummaydisNisikresistancegenesRhm1andRhm2 ,maizedwarfmosaicvirusresis tancegeneMdm1,wheatstreakmosaicvi…     

Chromosome G-banding in situ hybridization of RFLP marker umc58 linked with the gene hm1 dictatingHelminthosporium carbonum susceptibility1 in maize

The technique of simultaneous G-banding and in situ hybridization has been developed in plants for the first time. Using this technique, RFLP marker umc58 closely linked with the hm1 gene dictatingHelminthosporium carbonum susceptibility1 was localized onto 1L3 (chromosome 1, long arm, the third band from the centromere to the end of the arm), 5L5 and 9L5. The results demonstrated that umc58 was a triplicated sequence. It was deduced that umc58 probably was in a duplicated region that includes a part ofHelminthosporium carbonum susceptibility genes (hm1 and hm2), as the hybridization sites of umc58 in chromosomes 1 and 9 were those at which the genes localize. The techniques of simultaneous G-banding and ISH in plants are discussed.     

C-banding pattern and nucleolar organizer regions of Cynoglossus semilaevis Günther, 1873

The C-banding pattern and nucleolar organizer regions of the metaphase chromosomes of Cynoglossus semilaevis are reported. The interstitial regions in all chromosomes including the pair of sex chromosomes had positive C-bands, and the 6th and 12th pairs of chromosomes were entirely stained in most cases. Some chromosomes such as the 1st, 2nd, 7th, 8th, 9th and 10th pairs showed C-bands at centromeric or distal ends. The C-banding heretochromatin occupies 30.03% of the total chromosome surface in C. semilaevis, which is similar to that of amphibians such as Mixophyes fasciolatus (30.2%) and M. schevilli (20.7%), but is rather lower than that of cephalochordate Branchiostoma belcheri (54.3%). Silver staining revealed a single pair of nucleolar organizer regions (NORs) located in the telomeric regions of Chromosome 2. The association of NORs with heterochromatin observed in vertebrates also occurs in C. semilaevis as the telomeric regions of Chromosome 2 are always stained positively with C-banding.     

C-banding pattern and nucleolar organizer regions of Cynoglossus semilaevis Günther, 1873

The C-banding pattern and nucleolar organizer regions of the metaphase chromosomes of Cynoglossus semilaevis are reported. The interstitial regions in all chromosomes including the pair of sex chromosomes had positive C-bands, and the 6th and 12th pairs of chromosomes were entirely stained in most cases. Some chromosomes such as the 1st, 2nd, 7th, 8th, 9th and 10th pairs showed C-bands at centromeric or distal ends. The C-banding heretochromatin occupies 30.03% of the total chromosome surface in C. semilaevis, which is similar to that of amphibians such as Mixophyes fasciolatus (30.2%) and M. schevilli (20.7%), but is rather lower than that of cephalochordate Branchiostoma belcheri (54.3%). Silver staining revealed a single pair of nucleolar organizer regions (NORs) located in the telomeric regions of Chromosome 2. The association of NORs with heterochromatin observed in vertebrates also occurs in C. semilaevis as the telomeric regions of Chromosome 2 are always stained positively with C-banding.     

FISH analysis of the integration patterns in transgenic rice co-transformed by microprojectile bombardment

Using multi-color fluorescencein situ hybridization (FISH), we localized transferredbarnase-ps1 andpHctinG DNA sequences onto chromosomes of two transgenic rice plants, named Q12 and Q13, both of which were produced by micro-projectile bombardment. In both Q12 and Q13, each detected cell showed 2–3 signal spots on their chromosomes respectively. The signals of bothbarnase-ps1 andpHctinG were mostly detected in the adjacent chromosomal sites in which their signals were overlapped and could be recognized by the signal color on the metaphase chromosomes. Fiber FISH further demonstrated that the multiple copies in each of the two DNA sequences distributed adjacently on the DNA fiber in Q13. Combined with the results of Southern hybridization, the possible integration patterns in transgenic rice co-transformed by micro-projectile bombardment have been discussed.     

Development and characterization of interspecific hybrids between <Emphasis Type="Italic">Oryza sativa</Emphasis> and <Subscript>O. latifolia</Subscript> by <Subscript>in situ</Subscript> hybridization

Oryza sativa and O. latifolia belong to the AA and CCDD genomes of Oryza, respectively. In this study, interspecific hybrids of these species were obtained using the embryo rescue technique. Hybrid panicle traits, such as long awns, small grain, exoteric large purple stigma, grain shattering and dispersed panicles, resemble that of the paternal parent, O. latifolia, whereas there is obvious heterosis in such respects as plant height, tillering ability and vegetative vigor. Chromosome pairing and the genomic components of the hybrid were subsequently investigated using genomic in situ hybridization （GISH） and fluorescent in situ hybridization （FISH） analysis. Based on the mitotic metaphase chromosome numbers of the root tips investigated, the hybrid is a triploid with 36 chromosomes. The genomic constitution of the hybrid is ACD. In the meiotic metaphase Ⅰ of the hybrid pollen mother cell, poor chromosome pairing was identified and most of the chromosomes were univalent, which resulted in complete male sterility in the hybrid.     

Distribution of highly repeated DNA sequences in Haynaldia villosa and its application in the identification of alien chromatin

Haynaldia villosa (L.) is a wild relative species of common wheat that possesses many beneficial genes that can be used for wheat improvement. The accurate detection of H. villosa chromosomes in the genetic background of wheat is critical for transferring its beneficial genes to common wheat by chromosome engineering. The aim of the present study was to investigate the distribution patterns of two repeated DNA sequences, pSc119.2 and pAs1, as well as two rDNA multigene family sequences, 45S rDNA and 5S rDNA, in the individual chromosomes of H. villosa for the future precise identification of alien chromatin in germplasm development and breeding programs. A set of common wheat-H. villosa disomic addition 1V-7V lines was used to determine these specific signals on individual chromosomes of H. villosa. The results showed that two rDNA probes, pTa71 (45S rDNA) and pTa794 (5S rDNA), were located on 1VS and 5VS, respectively, and the signal could be discriminated exclusively in the common wheat background as effective markers of 1VS and 5VS. Furthermore, all seven chromosomes of H. villosa could be distinguished clearly by fluorescence in situ hybridization using pSc119.2 and pAs1 as probes in combination. The utilization of these cytogenetic markers of repetitive sequences, combined with other molecular markers sometimes, will make it possible for a precise identification of alien chromosomes with high efficiency.     

Genetic imprinting suggested by maternal heterodisomy in nondeletion Prader-Willi syndrome

Prader-Willi syndrome (PWS) is the most common form of dysmorphic genetic obesity associated with mental retardation. About 60% of cases have a cytological deletion of chromosome 15q11q13 (refs 2, 3). These deletions occur de novo exclusively on the paternal chromosome. By contrast, Angelman syndrome (AS) is a very different clinical disorder and is also associated with deletions of region 15q11q13 (refs 6-8), indistinguishable from those in PWS except that they occur de novo on the maternal chromosome. The parental origin of the affected chromosomes 15 in these disorders could, therefore, be a contributory factor in determining their clinical phenotypes. We have now used cloned DNA markers specific for the 15q11q13 subregion to determine the parental origin of chromosome 15 in PWS individuals not having cytogenetic deletions; these individuals account for almost all of the remaining 40% of PWS cases. Probands in two families displayed maternal uniparental disomy for chromosome 15q11q13. This is the first demonstration that maternal heterodisomy--the presence of two different chromosome 15s derived from the mother--can be associated with a human genetic disease. The absence of a paternal contribution of genes in region 15q11q13, as found in PWS deletion cases, rather than a mutation in a specific gene(s) in this region may result in expression of the clinical phenotype. Thus, we conclude that a gene or genes in region 15q11q13 must be inherited from each parent for normal human development.     

Tracing the location of powdery mildew resistance-related gene Stpk-V by FISH with a TAC clone in Triticum aestivum-Haynaldia villosa alien chromosome lines

Bacterial artificial chromosomes(BACs)or yeast artificial chromosomes(YACs)containing large inserts as probes for fluorescence in situ hybridization(FISH)have been used in the physical mapping of specific DNA sequences,especially for single-or low-copy sequences.Our earlier study identified Stpk-V,a powdery mildew resistance-related gene located on the 6VS chromosome arm of the wild grass Haynaldia villosa(tribe Triticeae),and obtained several Triticum aestivum–H.villosa alien chromosome lines carrying the Stpk-V gene.However,the precise physical location of the Stpk-V gene on chromosome 6VS is not known.In this study,we used TAC-FISH with TAC15 as the probe coupled with sequential genomic in situ hybridization(GISH)to determine the physical location of the Stpk-V gene in different T.aestivum–H.villosa 6V alien chromosome lines,including addition,substitution and translocation lines.The result indicated that the fraction length of the Stpk-V locus is 0.575±0.035 on the 6V chromosome short arm and this was confirmed by FISH using TAC15 as the probe for tracing the Stpk-V gene in other genetic stocks.The cytological mapping strategies used in this study will be of benefit for tracing the alien gene location in the course of introducing desirable traits from wild species.     

Sequence and analysis of chromosome 1 of the plant Arabidopsis thaliana

The genome of the flowering plant Arabidopsis thaliana has five chromosomes. Here we report the sequence of the largest, chromosome 1, in two contigs of around 14.2 and 14.6 megabases. The contigs extend from the telomeres to the centromeric borders, regions rich in transposons, retrotransposons and repetitive elements such as the 180-base-pair repeat. The chromosome represents 25% of the genome and contains about 6,850 open reading frames, 236 transfer RNAs (tRNAs) and 12 small nuclear RNAs. There are two clusters of tRNA genes at different places on the chromosome. One consists of 27 tRNA(Pro) genes and the other contains 27 tandem repeats of tRNA(Tyr)-tRNA(Tyr)-tRNA(Ser) genes. Chromosome 1 contains about 300 gene families with clustered duplications. There are also many repeat elements, representing 8% of the sequence.     

Centromere inactivation in a dicentric rice chromosome during sexual reproduction

T0135 is a variant selected from the progeny of a rice line telotrisomic for the short arm of chromosome 11 （2n＋IIS＇）. Fluores- cent in situ hybridization （FISH） results indicated that T0135 contained two telocentric chromosomes, which have two centro- mere-specific molecular markers （5S rDNA） for chromosome 11; thus T0135 is a newly-described rice chromosome variant with two dicentric chromosomes, named 22＋11L-＋11L＇＋I IS.11S-＋I 1S-11S. （22 represents the 22 chromosomes excluding chromo- some 11 in the rice genome, ＂-＂ represents the centromere）. To investigate the genetic stability of the rice dicentric chromosomes during sexual reproduction, we observed the chromosome types in the progeny. Ninety-four percent of the progeny had the same chromosome type as the parental line. This result indicates that the dicentric chromosomes are mostly stable during mitosis and meiosis. Immunofluorescence analysis for centromere specific histone H3 （CENH3） revealed that only one centromere is active and the other centromere is inactivated in the rice dicentric chromosomes.     

大豆bHLH转录因子家族成员的进化及功能分化研究

碱性螺旋-环-螺旋蛋白(basic Helix-Loop-Hleix,bHLH)转录因子家族是动植物中最大的转录因子家族之一,主要由碱性氨基酸区域和螺旋-环-螺旋区域组成,在动植物生长发育和胁迫应答反应中发挥着重要作用.本研究通过对大豆全基因组生物信息学分析和分子生物学研究手段,深入研究了大豆bHLH基因家族的进化机制,同时探讨了该基因家族在大豆中的功能分化.结果表明,大豆中含有340个非冗余的bHLH基因家族成员,通过对这些成员的Pfam蛋白结构域、Motif组成和系统进化关系的分析,将这些成员分为15组共24个亚家族,在大豆20条染色体上呈现不均匀分布.bHLH理化性质差异较大,编码的大豆氨基酸数量为91～815aa,相对分子量为10 273.88～91 300.38,理论等电点为4.56～10.40;碱性氨基酸区含有His5-Glu9-Arg13保守序列,与靶基因结合有关,HLH区含有Arg23和Arg55,与形成二聚体有关,同时含有5种保守元件.多数成员在大豆根以及花发育的5个时期中具有组织表达特异性,不同基因表达量差异较大.     

The DNA sequence and analysis of human chromosome 13

Chromosome 13 is the largest acrocentric human chromosome. It carries genes involved in cancer including the breast cancer type 2 (BRCA2) and retinoblastoma (RB1) genes, is frequently rearranged in B-cell chronic lymphocytic leukaemia, and contains the DAOA locus associated with bipolar disorder and schizophrenia. We describe completion and analysis of 95.5 megabases (Mb) of sequence from chromosome 13, which contains 633 genes and 296 pseudogenes. We estimate that more than 95.4% of the protein-coding genes of this chromosome have been identified, on the basis of comparison with other vertebrate genome sequences. Additionally, 105 putative non-coding RNA genes were found. Chromosome 13 has one of the lowest gene densities (6.5 genes per Mb) among human chromosomes, and contains a central region of 38 Mb where the gene density drops to only 3.1 genes per Mb.     

FISH of 5S rDNA and telomeric (TTAGGG) n repeats in normal and translocated populations of the frog Quasipaa boulengeri (Anura, Ranidae)

Quasipaa boulengeri,a spiny frog,is widely distributed in the low mountain regions,around Sichuan Basin.Our previous study revealed five karyotypes,caused by a translocation,that are randomly distributed throughout different populations.5S rDNA and telomere sequence(TTAGGG) n are potential good markers for chromosome identification and karyological evolution.In this study,we examined the sequences of 14 populations using fluorescence in situ hybridization(FISH) to detect if there is any variation between karyologically normal and translocated populations.5S rDNA loci were located at the same position on chromosomes 1 in 7 translocated populations.In two of the seven normal populations,5S rDNA also occurred on chromosome 5 in addition to chromosome 1.Our findings further indicate that the 5S rDNA on No.1 most likely represents the ancestral condition,while the minor loci represent the derived state.Signal density variations of the 5S rDNA were observed beteween homologous chromosomes or sister chromatids of pair 1 in both normal and translocated populations.Telomere sequences were identically located on all ends of the 26 chromosomes in seven rearranged populations,however,no ITSs were observed on the translocated chromosomes 1 and 6.Two of the six normal populations were found to contain ITSs which indicates that populations with translocation events diverged prior to those with ITSs rearrangements.In the KKS and BF populations,the ITSs of chromosome 3 are not always found on both homologues.Inter-chromosomal signal strength of telomeric sequences commonly differs within all populations.     

Clustering of breakpoints on chromosome 11 in human B-cell neoplasms with the t(11;14) chromosome translocation

The t(11;14) (q13;q32) chromosome translocation has been reported in diffuse small and large cell lymphomas and in chronic lymphocytic leukaemia (B-CLL) and multiple myeloma. Because chromosome band 14q32 is involved in this translocation, as well as in the t(8;14) (q24;q32) translocation of the Burkitt tumour, interruption of the immunoglobulin heavy-chain locus was postulated for this rearrangement. We have cloned the chromosomal joinings between chromosomes 11 and 14 and also between chromosomes 14 and 18, in B-cell tumours carrying translocations involving these chromosomes, and suggested the existence of two translocated loci, bcl-1 and bcl-2, normally located on chromosomes 11 (band q13) and 18 (band q21) respectively, involved in the pathogenesis of human B-cell neoplasms. The results indicate that in the leukaemic cells from two different cases of CLL, the breakpoints on chromosome 11 are within 8 nucleotides of each other and on chromosome 14 involve the J4-DNA segment. Because we detected a 7mer-9mer signal-like sequence with a 12-base-long spacer on the normal chromosome 11, close to the breakpoint, we speculate that the t(11;14) chromosome translocation in CLL may be sequence specific and may involve the recombination system for immunoglobulin gene segment (V-D-J) joining.     

Construction of a bacterial artificial chromosome library for Gossypium herbaceum var. africanum

Gossypium herbaceum var. africanum is the only wild cotton species within the cultivated diploid G. herbaceum. As the A sub-genome donor of tetraploid cotton, it is characterized by its resistance to insects, diseases, and other adversities. We have constructed the first bacterial artificial chromosome library (BAC) for G. herbaceum var. africanum. With high quality and broad coverage, this library includes 75000 clones, with an average insert size of about 115 kb and fewer than 4% empty clones. Our library is approximately five-fold the size of the A-genome (1667 Mb) and it provides 99.3% probability for isolating genes of interest or their sequences. Using nine SSR markers that are located on five different chromosomes and linked with resistance to Verticillium wilt, seven of nine could amplify the 40 superpools and got 1-14 hits. Because of its moderate wide coverage and relative large insert size, this library will be an important genomic resource for classifying and analyzing the evolution of cotton species, as well as for isolating disease-resistance genes and control elements.     

临汾附近两种蛙类染色体特征研究

本文利用骨髓细胞空气干燥法，对临汾附近两种蛙类染色体组型进行分析．其中中国林蛙(Rana chensinensis)的二倍体染色体数目为2n＝24，包括12个大型染色体(相对长度大于7．0)和12个小型染色体(相对长度小于6，0)．全部染色体可配成12对．黑斑侧褶蛙(Pelophylax nigromaculata)的二部体染色体数目为2m＝26，全部染色体可配成13对，5对大型、8对小型染色体．与北京郊区同种蛙类染色体比较，说明不同地理居群的同种蛙染色体具有丰富的多样性．     

Development of specific chromosomal DNA pool for rice field eel and their application to gene mapping

The chromosomes 1, 3, 5, 6, 7, 10 and 12 of rice field eel (Monopterus albus Zuiew) have been microdissected successfully from meiosis Ⅰ diakinesis spreads by using glass microneedle under an inverted microscope. And the DOP-PCR products of the single chromosome dotted on the nylon membrane as “specific chromosomal DNA pool”, have been hybridized with 6 probes to map these genes. The mapping results show that Zfa has been mapped to chromosome 1, rDNA to chromosomes 3 and 7, both Gh and Pdegg to chromosome 10, Hsl to chromosome 5 and Hox genes have been detected on chromosomes 1, 3, 6 and 10 meantime. It has initiatively been suggested that chromosome 10 of rice field eel might possess the commom conserved synteny to that on chromosome 17 of human, chromosome 11 of mouse, chromosome 12 of pig and chromosome 19 of bovine. And so chromosome 3 of rice field eel might also contain the commom conserved synteny to that on chromosome 2 of zebrafish. Our study is an attempt to establish a new and feasible method to advance the study of gene mapping and chromosome evolution in fish, and also to provide a new idea to distinguish each chromosome on the base of molecular markers for fish.