Assignment of unanchored scaffolds in genome of Brassica napus by RNA-seq analysis in a complete set of Brassica rapa-Brassica oleracea monosomic addition lines

The economically valuable oil crop Brassica napus(AACC, 2 n=38), which arose from interspecific hybridization between the diploid ancestors Brassica rapa(AA, 2 n=20) and Brassica oleracea(CC, 2 n=18), has a complex genome. More than 10% of the assembled sequences, most of which belong to the C subgenome, have not been anchored to the corresponding chromosome. Previously, a complete set of monosomic alien addition lines(MAALs, C1–C9) with each of the nine C-subgenome chromosomes added to the extracted A subgenome was obtained from the allotetraploid B. napus donor Oro, after the ancestral B. rapa(RBR Oro) genome was restored. These MAALs effectively reduced the complexity of the B. napus genome. Here, we determined the expression values of genes on unanchored scaffolds in the MAALs and RBR Oro. Then, multiple comparisons of these gene expression values were used to determine the affiliations of the nonanchored scaffolds on which the genes were located. In total, 54.68%(44.11 Mb) of the 80.67 Mb of non-anchored scaffolds belonging to the C subgenome were assigned to corresponding C chromosomes. This work highlights the potential value of these MAALs in improving the genome quality of B. napus.     

Obtaining and Cytological Identification of a Set of Primary Trisomics in Cabbage

Selection of primary trisomics of the cabbage （Brassica oleracea var.capitata L） forms an important basis for gene chromosome mapping and for other genetic studies. The cabbage self-fertilization line - 9601 was used as material, using the root-tip cell chromosome number and pollen mother cell chromosome number identification and karyotype analysis to select the primary trisomics from the progenies of 3x x 2x in the cabbage. Many aneuploid plants with one or two extra chromosomes were obtained and a set of primary trisomics （Tri-1, Tri-2, Tri-3, Tri-4, Tri-5, Tri-6, Tri-7, Tri-8, and Tri-9, in which the Tri-1 and Tri-4 were from 2n＋2 plants and others from 2n＋ 1 plants） was acquired from these plants. Each trisomic exhibited some unique features, such as plant height, plant type, leaf type, size of flower bud, and inflorescence. The triploid crossing by the diploid is a convenient and effective way to select trisomics in the cabbage.     

Genetic Analysis and Preliminary Mapping of a Highly Male-Sterile Gene in Foxtail Millet（Setaria italica L.Beauv.） Using SSR Markers

Breeding of male-sterile lines has become the mainstream for the heterosis utilization in foxtail millet,but the genetic basis of most male-sterile lines used for the hybrid is still an area to be elucidated.In this study,a highly male-sterile line Gao146A was investigated.Genetic analysis indicated that the highly male-sterile phenotype was controlled by a single recessive gene a single recessive gene.Using F 2 population derived from cross Gao146A/K103,one gene controlling the highly male- sterility,tentatively named as ms1,which linked to SSR marker b234 with genetic distance of 16.7 cM,was mapped on the chromosome VI.These results not only laid the foundation for fine mapping of this highly male-sterile gene,but also helped to accelerate the improvement of highly male-sterile lines by using molecular marker assisted breeding method.     

Characterization of RAPD Markers, and the RFLP Marker Linked to Powdery Mildew Resistance Gene Derived from Different Accessions of H. villosa

The analysis was carried out on performance of the resistance gene from Haynaldia villosa accession of the former Soviet Union to different isolates ofBluemerie graminis. Polymorphisms were revealed between 6D/6V substitution line Pm930640 and its pedigree parents using five RAPD markers of OPAN031700, OPAI01700, OPAL03750, OPAD07480 and OPAG 15580 screened out from 120 random 10-mers primers. Three RAPD markers of OPAN03, OPAI01 and OPAL03 were linked with the resistance gene by analysis of F2 population of ChancellorxPm930640. Analysis of 29 wheat lines including part of lines conferring the known genes from Pro1 to Pro20 respectively, lines conferring resistance gene from two H. villosa accessions and the related wheat parents, were analyzed and the results showed that these markers not only linked to the gene resistant to powdery mildew from H. villosa, but also detected different genetic backgrounds. OPAL03750 can be used as the marker to distinguish the different resistant lines from two H. villosa accessions because it was only observed in the materials from H. villosa of the former Soviet Union. RFLP analysis also showed the polymorphisms between two H. villosa accessions and their derived resistant lines.     

Cytological Evaluation and Karyotype Analysis in Plant Germplasms of Elytngia Desv.

Elytrigia Desv. is widely distributed throughout the world and is represented with species of various levels of ploidy including diploids, tetraploids, hexaploids, octaploids, and decaploids. The distribution pattern of these ploidy levels, however, is not well-defined. In this study, the levels of ploidy for 64 accessions of Elytrigia from 25 countries were determined with microscopic procedures. The results showed that accessions of E. intermedia and E. repens were grouped into three distinct levels of ploidy including diploids, tetraploids and hexaploids. For E. elongata, E. pontica, and E. caespitosa, it was found that two ploidy levels presented, and only one ploidy level was in those of E. hybrid, E. pycnantha, E. pungens, E. juncea, and E. alatavica. Karyotype analysis indicated that the karyotype formula of diploid, tetraploid and hexaploid of E. intermedia was 2n = 2x = 14 = 6m ＋ 6sm ＋ 2st, 2n = 4x = 28 = 2M ＋ 10m ＋ 16sm and 2n = 6x = 42 = 4M ＋ 18m ＋ 20sin, respectively. Furthermore, the karyotype formula of three germplasms in tetraploid of E. intermedia was 2n=4x=28 =2M＋ 10m＋ 16sm, 2n=4x=28=4M＋22m （sat）＋2sm and 2n=4x=28 =4M＋ 12m＋ 12sm （sat）, which were not completely uniform. Therefore, it could be suggested that the studies about chromosome constitution would be helpful for the detail understanding of the diversity of germplasm resource in Elytrigia and promoting the utilization in the crop molecular breeding.     

Diallel Crossing Analyses of Resistance to Main Diseases in Pepper （Capsicum annuum L.）

Fifteen capsicum combinations were made with 6 parents by （1/2）n（n-1） diallel crossing. Genetic parameters in the resistance to TMV, CMV, phytophthora blight, bacterial spot of these combinations were studied by Hayman. The results indicated that the resistance to TMV, CMV and bacterial spot conformed genetically to the “additive-dominant” model but the resistance to phytophthora blight did not and significant epistatic dominance effect existed in it. F1 hybrid＇s resistance to CMV was controlled by homozygous dominant gene （s）, but resistance to bacterial spot by heterozygous one （s）. There were little, or no sum of dominant effect and genomes controlling the dominant expression of F1 hybrids in its phytophthora blight resistance.     

Introduction of herbicide resistant gene （bar） into maize （Zea mays L.） via pollen tube pathway

The pollen tube pathway method of transformation has been reported to be successful in most crops,but less successfu in maize.DNA can be transferred by cutting the stigma following pollination and applying the DNA solution in a suitable period DNA presumably reaches the ovary by flowing down the pollen tube and then integrates into the just fertilized but undivided zygotic cells.To provide the molecular evidence for this procedure,the plasmids pGBIRC carrying a CaMV35S promoter-PPT acetyle transferase(bar)gene-nos terminator gene fusion construct were used.Total 3 276 seeds were produced from the ears treated with DNA.It was found that 35 seedlings were GUS assay positive,but less intense than that of the positive controls,of which 17 were PCR amplification positive.But,only 13 of the seeds from the plants treated with DNA containing the bar gene were found to be resistant compared with the negative control.Less than 1.07% of progeny seedlings tested expressed a herbicide positive reaction and polymerase chain reaction(PCR)with seedling DNA did detect the bar gene.Morphological variation was observed in six plants.We succeed in obtain PPT-resistant maize inbred lines via pollen tube pathway.     

Analysis of Disequilibrium Hybridization in Hybrid and Backcross Progenies of Saccharum officinarum × Erianthus arundinaceus

Erianthus arundinaceus is an important, closely related genus of Saccharum officinarum L. It is therefore important to understand how the chromosomes are transmitted when it hybridizes with sugarcane. The hybrids and backcross progenies of S. officinarum and E. arundinaceus and their parents were used for Karyotype analysis and to study the law of chromosome transmission. The results showed that the somatic chromosome number of both of the E. arundinaceus Hainan92-105 and Hainan92-77 were 2n = 60 = 60sm, belonging to type 1 A, and the BC1 YC01-21 was 2n = 104 = 100m ＋ 4sm, belonging to type 2C. The other six tested clones belonged to type 2B. The both F1s YC96-66 and YC96-40 that originated from Badila （2n = 80 = 70m ＋ 10sm） with E. Arundinaceus were 2n = 70 = 68m ＋ 2sm, which suggests an n ＋ n transmission. The cross between YC96-66 （female parent） and CP84-1198 （male parent, 2n = 120 = 114m ＋ 6sm） also followed the same genetic law and the somatic chromosome number of their progeny, YC01-3 （2n = 105 = 95m ＋ 10sm）. The cross derived from YC96- 40 （female） and CP84-1198 （male）, YC01-21 had 2n = 104 = 100m ＋ 4sm chromosomes, following the same genetic law of n ＋ n. However, YC01-36 had 2n = 132 = 130m ＋ 2sm chromosomes, which suggests a 2n ＋ n chromosome transmission. It can be inferred that the inheritance of chromosomes was very complex in the BC1. The difference in chromosome number between clones was as high as 28. This could be explained by the 2n ＋ n transmission of chromosomes. In addition, as there was not be a regular number of haploids, this phenomenon is termed as disequilibrium hybridization.     

Genetic Diversity of Recurrent Selection Populations with Ms2 Gene Assessed by Gliadins in Common Wheat （Triticum aestivum L.）

The male-sterile lines with Ms2 gene were highly evaluated in recurrent selection in wheat （Triticum aestivum L.）. Three populations C6 （population after six cycles of selection）, C7 （population after seven cycles of selection）, and C8 （population after eight cycles of selection） were constructed through recurrent selection with 12 parental materials （P）. Acid polyacrymide gel electrophoresis （A-PAGE） analysis was used to identify gliadin patterns and evaluate the genetic diversity in 12 parents and three populations. A total of 63 bands were identified, of which 17 polymorphic bands and 7 unique bands were present in populations and seven polymorphic bands and four unique bands were present in parents. The number of polymorphic and unique bands decreased gradually from C6 to C8, especially for to- and y-gliadins. The genetic distances in C6, C7, and C8 were calculated. The distributions of genetic distance were different in three recurrent selection populations. From C6 to C8, the genetic distance was 0.2687, 0.2652 and 0.1987, respectively. Statistically significant differences were detected between C7 and C8 with the T value of 37.9718. The result of cluster analysis based on genetic similarity matrix of three populations fitted well to those of principle coordinates analysis （PCoA）. Compared with 12 parents, almost all individuals of three populations are new genotypes. Most of the individuals from C6 and C7 could be divided into two groups, while most individuals of C8 were in one cluster. In conclusion, the results indicated that the genetic diversity was decreased severely according to the information revealed by A-PAGE, although some variations could be created in the recurrent selection. It was necessary to introduce diverse germplasm based on the genetic database of recurrent population to maintain and improve the breeding efficiency in the further program.     

Paternity Testing of Selected Abaca （Musa textilis L. Nee） Hybrids Using Morphometric Markers

Investigation into the paternity of four abaca （Musa textilis L, Nee） hybrids was done to ascertain the mode of transmission of selected morpho-agronomic traits and to detect possible heterosis. In situ morphological characterization was undertaken using twenty five qualitative and six quantitative characters. Results revealed that a great majority of the qualitative traits were shared by both parents and their hybrids. For the rest, the qualitative traits were inherited from one or the other parent though some variant phenotypes （i.e. chimerism） were also noted in the hybrids. Cases ofheterosis were also observed and this could be exploited to increase fiber yield in the hybrids. Though inconclusive due to factors such as the heterogenous nature of abaca plants in the field and the susceptibility of morphological traits to environmental fluctuations, this study has provided baseline information on abaca hybridity that can be verified using more robust technologies as molecular markers.     

Monosomic Addition Lines of Flowering Chinese Cabbage (B. campestris L. ssp. chinensis var. parachinensis L. H. Bailey)-Chinese Kale (B. oleracea L. var. alboglabra L. H. Bailey)

Interspecific alien addition lines have played significant roles in gene mapping, intergenomic gene transfer and chromosomal homoeological identification between closely related species. Selection of alien addition lines was conducted by karyotype analysis and morphological observation with the reference of parents. Triploid interspecies hybrid (AAC, 2n=3x=29) was obtained from Brassica campestris ssp. chinensis var. parachinensis Qinglu 9601 (tetraploid, AAAA, 2n=4x=40)×B. oleracea vat. alboglabra Baihua 9705 (diploid, CC, 2n=2x=18) by immature hybrid embryo culture in vitro. Five different alien monosomic addition lines (AA C2, AA C3, AA C4, AA C6, AA C7) were obtained from the backcross progenies of AAC×AA. Each alien monosomic addition line has some specific morphological characters. It is feasible to obtain alien addition lines from the progenies of AAC×AA by karyotype analysis and morphological observation based on the reference of parents.     

Generation and identification of four new monosomic addition lines of flowering Chinese cabbage-Chinese kale

A selection test of four new flowering Chinese cabbage (Brassica campestris L. var. utilis Tsen et Lee)-Chinese kale (Brassica alboglabra Bailey) monosomic alien addition lines was conducted by karyotype analysis and morphological marker identification. The results mainly showed the following: 61??2n + 1?? plants screened out from 655 progeny plants of the trigenomic hybrid (AAC) backcrossed with its parent flowering Chinese cabbage (AA), four new monosomic alien addition lines(AA + C₁, AA+ C₅, AA+ C₈ and AA + C₉) obtained from the 61 ??2n + 1?? plants, the transmission rates of the monosomic alien addition lines (AA + C₁, AA+C₅ and AA + C₈) of 19.7%, 23.44%, and 36.51% by female gametes and 7.84%, 8.89%, and 9.43% by male gametes, respectively, and the alien chromosome C-1 and C-5 had partial homology with at least one chromosome of A-genome.     

大白菜-结球甘蓝双单体异附加系的SSR鉴定及其特性

 【目的】筛选大白菜-结球甘蓝异附加系,利用SSR标记鉴定附加的外源染色体,并对异附加系特性进行研究。【方法】以大白菜-结球甘蓝异源三倍体（AAC）与二倍体大白菜（AA）回交一代的自交后代为试材,利用根尖染色体计数、甘蓝连锁群特异SSR标记等方法,筛选、鉴定大白菜-结球甘蓝异附加系。【结果】在BC1F2中筛选到2n=22的植株;该植株对位于甘蓝01连锁群的3对SSR引物和08连锁群的2对SSR引物的扩增结果与异源三倍体杂交种是一致的,而对位于其它7个连锁群的20对特异引物的扩增结果与大白菜一致,表明该植株附加的外源染色体与甘蓝的01、08连锁群相对应,为双单体异附加系;该双单体异附加系绝大多数花粉母细胞减数分裂终变期染色体构型为10Ⅱ+2Ⅰ,后期Ⅰ以11-11、10-12分离方式为主,后期Ⅱ产生了n=11配子。【结论】利用甘蓝连锁群相对于大白菜特异的SSR标记可以准确鉴定大白菜-结球甘蓝异附加系;获得了附加甘蓝01、08连锁群的大白菜双单体异附加系,为进一步合成附加甘蓝01或08连锁群大白菜-结球甘蓝单体和双体异附加系奠定基础。     

利用PCR技术初步鉴定小麦-加州野大麦异染色体系

为快速鉴定普通小麦与普通小麦—加州野大麦双二倍体杂交、回交后代植株的染色体组成，研究小麦背景中添加的外源染色体与小麦染色体之间的部分同源关系，选用已被定位在小麦7个部分同源群21条染色体上的38个SSR引物对杂种回交后代植株进行PCR扩增。结果表明，其中27个小麦SSR引物在普通小麦与小麦—加州野大麦双二倍体间有多态性扩增，涉及4个部分同源群的11对引物，可在不同杂种回交植株中扩增出与双二倍体相同的多态带纹；根据PCR扩增和细胞遗传学分析的结果，在18个回交后代中初步鉴定出7个可能的异附加系，其中2个二体异附加系、1个端二体异附加系、2个单体异附加系和2个双单体异附加系。所选育的异附加系分别涉及第1、2、4和7部分同源群。     

大白菜-结球甘蓝5号二体异附加系的选育及鉴定

【目的】获得稳定遗传的大白菜-结球甘蓝二体异附加系。【方法】以大白菜-结球甘蓝5号单体异附加系为材料,对其自交后代进行细胞学及SSR鉴定。【结果】37株自交后代中2n=22植株的比率为10.81%,通过对4个2n=22的植株进行核型分析及SSR鉴定,确定其为大白菜-结球甘蓝5号二体异附加系。减数分裂观察发现,大白菜-结球甘蓝5号二体异附加系后期Ⅰ虽然出现染色体落后现象,但在后期Ⅱ中染色体的分裂方式以11-11-11-11为主,其比率为69.55%。【结论】获得了大白菜-结球甘蓝5号二体异附加系,其外源染色体与甘蓝07连锁群相对应;该二体异附加系减数分裂形成的四分体以附加1条的染色体为主(n=11),有较高的遗传稳定性。二体异附加系的获得为研究芸薹属A、C基因组的亲缘关系,以及向大白菜中导入结球甘蓝的优良基因具有重要意义。     

线纹香茶菜狭基变种和线纹香茶菜细花变种的染色体核型分析

[目的]对线纹香茶菜狭基变种[Radbosia lophanthoides var.gerardianus(Benth.)Hara]和线纹香茶菜细花变种[Radbosia lophan-thoides Hara var.graciliflorus(Benth.)Hara]的染色体进行核型分析。[方法]分别对线纹香茶菜狭基变种和线纹香茶菜细花变种生长旺盛的茎尖进行染色,压片后对它们的染色体进行观察和分析。[结果]线纹香茶菜狭基变种的核型公式为K(2n)=2x=24=18m+4Sm,属2A型。线纹香茶菜细花变种的核型公式为K(2n)=2x=24=16m+8Sm,属1B型。[结论]线纹香茶菜细花变种比线纹香茶菜狭基变种更进比。     

以色列野生二粒小麦和光稃野燕麦杂交亲本与后代核型分析

 对以色列野生二粒小麦（母本）和光稃野燕麦（父本）远缘杂交亲本与后代的核型及进化关系进行了分析。结果表明：母本的染色体长度比为1626,核型公式为2n=4x=28=18m（2SAT）+10sm（2SAT）,核型为1A。父本的染色体长度比为2526,核型公式为2n=6x=42=20m（2SAT）+22sm（4SAT）,核型类型为2B。以色列野生二粒小麦和光稃野燕麦杂交后代0878株系的染色体数目为42,染色体长度比为1.802,核型公式为2n=6x=42=22m（2SAT）+20sm（2SAT）,核型为2A。 0878-1株系的染色体数目为42,染色体长度比为2.057,核型公式为2n=6x=42=38m+4sm（4SAT）,核型为2B。由于光稃野燕麦遗传物质渗入到该杂交后代,获得了进化程度高于其母本的普通小麦型小麦新种质。     

古田山黄精——多花黄精一新变种及其与近似种的比较研究

报道了浙江产黄精属植物一新变种--古田山黄精,并对其与近似种在形态学和核型上进行了比较研究.结果如下:古田山黄精(Polygonatum cyrtonema var.gutianshanicuym),2n=2χ=24=1 4m+4sm+6st(2 sc);多花黄精(P.cyrtonema),2n=2χ=22=10m+8sm+4st;长梗黄精(P.filipes),2n=2χ=16=8m(4sat)+8sm.长梗黄精的核型属于2B型;多花黄精属于3B型;古田山黄精属于2C型,核型均为首次报道.数据统计表明叶片宽度、总花梗长度是古田山黄精与其原变种多花黄精的主要区别所在.     

基于GISH的甘蔗与蔗茅属间杂交F1后代染色体组成及核型分析

 【目的】探讨甘蔗(Saccharum spp.)与蔗茅(Erianthus fulvus)杂交F1材料的染色体组成及核型差异。【方法】以甘蔗与蔗茅间杂交所获得的16份F1材料为研究对象进行单色及双色基因组原位杂交(genomic in situ hybridization,GISH)研究,在GISH的基础上对3份材料的中期染色体进行核型分析。【结果】GISH的研究结果表明,子代材料中具有7—10条不等的父本染色体;由于母本遗传物质对父本遗传物质的排斥作用引起的染色体消减及片段化导致父本染色体在子代中有断裂的现象;研究结果表明,在间期核中亲本染色质以分散的方式单独分布;3份材料的中期染色体进行核型分析的公式分别为2n=92=80m(4SAT)+12sm、2n=88=82m+6sm及2n=88=78m(2SAT)+10sm,核型分类为2C、2C和2B。【结论】甘蔗×蔗茅杂种F1的染色体组成为n+n及2n+n,它们的核型均为较原始的染色体类型。     

龙牙百合核型分析

采用染色体常规压片方法对龙牙百合根尖细胞进行了染色体核型分析,结果表明,龙牙百合的染色体数目为2n=24,染色体核型公式为2n =2x =24 =4m (SAT) +2sm+ 10st (2SAT) +8t,核型分类属于3B类型,核型不对称系数As.K为76.80％,染色体相对长度系数I.R.L=6L+4M2+10M1+4S.