Genome size variation in three Saccharum species

Saccharum species are autopolyploids with ploidy level ranging from 5× to 16x, and are considered the most complex genomes among crop plants. In present study, the genome sizes of 28 Saccharum spontaneum accessions, 15 Saccharum officinarum accessions, 28 Saccharum robustum accessions, and 12 Saccharum hybrids spp. were analyzed using flow cytometry. The estimated genome sizes of S. officinarum accessions ranged from 7.50 to 8.55?Gb with an average size of 7.88?Gb. In S. robustum, the estimated genome sizes ranged from 7.65 to 11.78, reflecting the variation of ploidy level. In S. spontaneum, the estimated genome sizes varied widely, with a range from 3.36 to 12.64?Gb, also due to variation of ploidy level. The average monoploid genome size of S. officinarum was 985?Mb, and that of S. spontaneum was 843?Mb. The results also showed that genome sizes were correlated with chromosome numbers, and based which, that the unknown chromosome numbers of some accessions could be predicted. The estimated genome sizes of Saccharum germplasm also helped identify some mislabeled accessions and yielded information critical for sugarcane breeding and genome sequencing programs.     

Genetic mapping of a PRSV-P resistance gene in “highland papaya” based on inheritance of RAF markers

Papaya ringspot virus type P (PRSV-P) is a significant disease of Carica papaya. A major gene for PRSV-P resistance has been mapped in Vasconcellea cundinamarcensis, a distant relative of C. papaya. This was achieved by genetic mapping of the resistance phenotype and inherited, dominant, polymorphic randomly amplified DNA fingerprint (RAF) markers in F2 progenies of V. parviflora and V. cundinamarcensis. The parents of this cross confer resistance to several major diseases that affect C. papaya including PRSV-P in V. cundinamarcensis. Heredity of DNA markers and PRSV-P resistance was studied in the intrageneric population presented due to intergeneric fertility barriers between Carica and Vasconcellea. Genetic polymorphism between parents, based on RAF markers, was 75% with more than 70% of markers generated showing mendelian segregation for the expected ratios 1:3 or 1:1 (p < 0.05). Preferential inheritance of markers from either parent was not detected in the F2, indicating stable transfer of the genetic material. Discrete V. parviflora and V. cundinamarcensis linkage maps were compiled from 79 and 83 framework markers, delineating to 10 and 11 groups respectively. F1 and F2 progeny were screened for resistance to PRSV-P under controlled conditions. The resistant phenotype segregated 3:1 in the F2 and mapped to V. cundinamarcensis linkage group 7 with adjacent RAF markers within 4 cM. The framework maps of V. parviflora and V. cundinamarcensis presented cover 630.2 and 745.4 cM respectively, accounting for between 47–52 and 49–55 percent of the predicted genome lengths. These maps provide a platform for further genetic study of disease resistance characteristics identified in these species and the development of DNA markers tightly linked to these traits, which could be applied to the breeding of resistant C. papaya cultivars.     

流式细胞术测定5种苔藓植物的基因组大小

作为藻类和维管植物之间的过渡类群,苔藓植物不仅占有重要的进化地位,它同时可作为园艺上的观赏植物。目前对很多苔藓植物的培养条件、基因组大小等信息非常缺乏,且已进行基因组测序的苔藓物种较少,这些都严重制约了苔藓植物系统学的研究和资源利用。本研究以基因组大小为942 Mb的野生番茄(Solanum pennellii)为内标,采用流式细胞术对5种苔藓植物的基因组大小进行测定,结果表明,拟短月藓(Brachymeniopsis gymnostoma)的基因组大小约为391.40 Mb,绒叶青藓(Brachythecium velutinum)的基因组大小约为386.96 Mb,尖叶梨蒴藓(Entosthodon wichurae)的基因组大小约为441.01 Mb,桧叶白发藓(Leucobryum juniperoideum)的基因组大小约为493.21Mb,明叶藓(Vesicularia montagnei)的基因组大小约为337.94Mb。本研究为苔藓植物的全基因组测序、分子生物学研究、亲缘进化研究以及园艺苔藓品种开发和育种等方面提供重要信息。     

Chromosome-level de novo genome assemblies of over 100 plant species

Genome sequence analysis in higher plants began with the whole-genome sequencing of Arabidopsis thaliana. Owing to the great advances in sequencing technologies, also known as next-generation sequencing (NGS) technologies, genomes of more than 400 plant species have been sequenced to date. Long-read sequencing technologies, together with sequence scaffolding methods, have enabled the synthesis of chromosome-level de novo genome sequence assemblies, which has further allowed comparative analysis of the structural features of multiple plant genomes, thus elucidating the evolutionary history of plants. However, the quality of the assembled chromosome-level sequences varies among plant species. In this review, we summarize the status of chromosome-level assemblies of 114 plant species, with genome sizes ranging from 125 Mb to 16.9 Gb. While the average genome coverage of the assembled sequences reached up to 89.1%, the average coverage of chromosome-level pseudomolecules was 73.3%. Thus, further improvements in sequencing technologies and scaffolding, and data analysis methods, are required to establish gap-free telomere-to-telomere genome sequence assemblies. With the forthcoming new technologies, we are going to enter into a new genomics era where pan-genomics and the >1,000 or >1 million genomes’ project will be routine in higher plants.     

A SCAR marker for the sex types determination in Colombian genotypes of Carica papaya

Sex type determination in papaya (Carica papaya L.) is very important for crop improvement processes because it accelerates the identification of the fruitful plants. The use of molecular technology provides a quick and reliable identification of sex types in plantlets growing in seedbeds. Random amplified polymorphic DNA (RAPD) markers were used to determine the sex types of Colombian cultivars of dioecious papaya genotypes. This species has three sex types (male, female and hermaphrodite) determined by a multiallelic locus. There are no morphological differences at the chromosome level; therefore the identification of sex types by chromosomal dimorphism is not possible. A RAPD marker of 900 bp was found in male plants, but not in females or hermaphrodites. From this RAPD marker a sequence characterized amplified region (SCAR) was developed and it was possible to amplify fragments from the genomes of male and hermaphrodite plants, but not the female ones. The results indicate that this new SCAR marker will be valuable to determine the sex type of papaya plants.     

Comparative QTL mapping for male sterility of cultivated strawberry (Fragaria × ananassa Duch.) using different reference genome sequences

Male sterility is one of the reproductive isolation systems in plants and quite useful for F₁ seed production. We previously identified three independent quantitative trait loci (QTLs) for male sterility of cultivated strawberry, Here, we identified the specific subgenomes in which these QTLs are located by QTL-seq approach. QTLs qMS4.1, qMS4.2, and qMS4.3 were mapped separately in subgenomes Fvb4-4, Fvb4-3, and Fvb4-1, respectively, in ‘Camarosa’ genome assembly v. 1.0.a1. Candidate regions of qMS4.1 and qMS4.3 were clearly detected around 12–26 Mb in Fvb4-4 and 12–14 Mb in Fvb4-1, respectively; those of qMS4.2 were fragmented in Fvb4-3, which suggests that some scaffolds were incorrectly assembled in Fvb4-3. qMS4.3 was mapped to chr4X1 of ‘Reikou’ genome assembly r2.3, and qMS4.1 and qMS4.2 were both mapped to chr4Av, which indicates that differentiation of the subgenomes in which both QTLs are located was insufficient in ‘Reikou’ r2.3. Although ‘Camarosa’ genome assembly v. 1.0.a1 is an unphased map, which merges homologous chromosomes into one sequence, ‘Reikou’ genome assembly r2.3 is a phased map, which separates homologous chromosomes. QTL mapping to different reference genomes clearly showed the specific features of each reference genome, and that using different kinds of reference map could accelerate fine mapping and map-based cloning of certain genes of cultivated strawberry.     

Maternal inheritance of cytoplasmic organelles in intergeneric hybrids of Carica papaya L. and Vasconcellea spp. (Caricaceae Dumort., Brassicales)

The mode of inheritance of chloroplast and mitochondrial DNA has been determined in intergeneric hybrids between C. papaya and four different Vasconcellea species by employing a restriction fragment length polymorphism analysis of a PCR-amplified chloroplast and mitochondrial DNA region. Artificial F₁ hybrids were produced between a female specimen of C. papaya and male specimens of either V. parviflora, V. goudotiana, V. cundinamarcensis or V. quercifolia. The hybridization patterns of all hybrids correspond in all cases with that of the C. papaya mother, and are different from that of the paternal Vasconcellea species, thus indicating the maternal inheritance of cpDNA and mtDNA in intergeneric hybrids between C. papaya and wild relatives of the Vasconcellea genus.     

RFLP analysis of genetic variation in species of section Arachis, genus Arachis (Leguminosae)

Four A-genome species of the genus Arachis (A. cardenasii, A. correntina, A. duranensis, A. kempff-mercadoi), three B genomes species (A. batizocoi, A. ipaënsis and A. magna),the AABB allotetraploid A. hypogaea (cultivated peanut) and introgression lines resulting from a cross between A. hypogaea and A. cardenasii were analyzed by RFLP. The A genome species (cytologically characterized by the presence of a small chromosome pair ‘A’) were closely similar to each other and shared a large number of restriction fragments. In contrast, the B genome species differed more from one another and shared few fragments. The results of this study indicate that the absence of the small chromosome pair is not a good criterion for grouping species of section Arachis as B genome species, since their genome might be quite distinct from the B genome of A. hypogaea.The lowest genetic variation was detected within accessions of A. duranensis (17 accessions), followed by A. batizocoi (4 accessions) and A. cardenasii (9 plants of accession GKP 10017).The high level of genetic variation found in A. cardenasii might indicate that not all accessions of wild species of Arachis are autogamous, as reported for A. hypogaea.     

Application of DNA fingerprints for identification and genetic analysis of Carica papaya and other Carica species

Summary Various DNA fingerprint probes were applied to Carica papaya and other Carica species for both identification and genetic analysis.Each of the Carica papaya cultivars is characterized by a specific DNA fingerprints pattern. Various Carica species also have specific patterns which distinguish them from one another. Band sharing levels were used to estimate the relatedness between the various Carica species.Genetic analysis of 11 progeny from a cross between the Carica papaya cultivars 17/82 and 112 suggests that application of DNA fingerprinting to Carica papaya breeding, could make the process more efficient. Genetic analysis of the DNA fingerprint bands revealed no linkage or allelic relationship among the bands analyzed, indicating that these loci are not clustered in the Carica genome.     

Interspecific hybridization in <Emphasis Type="Italic">Sarcococca</Emphasis> supported by analysis of ploidy level,genome size and genetic relationships

Knowledge of ploidy level differences, genome size and genetic relationships between species facilitates interspecific hybridization in ornamentals. For Sarcococca (Buxaceae) only limited (cyto)genetic information is available. The aim of this study was to determine the genome size and chromosome number and to unravel the genetic relationships of a breeder’s collection using AFLP marker analysis. Based on these results, interspecific crosses were made and the efficiency and hybrid status was verified. Two groups of diploid plants (2n = 2x = 24) were observed, with either a genome size of 4.11–4.20 or 7.25–9.63 pg/2C. All the tetraploid genotypes (2n = 4x = 48) had genome sizes ranging from 7.91 to 8.18 pg/2C. In crosses between parents with equal ploidy level and genome size a higher crossing efficiency (on average 58% of the hybridizations resulting in fruits) and more true hybrids (on average 96% of the offspring) were obtained compared to crosses between plants with different genome size and ploidy level (on average 23% fruits and 24% hybrids, respectively). In none of the cross combinations, the ploidy level or genome size was found to be a complete hybridization barrier, although unilateral incongruity was found in some cross combinations. Distant genetic relationships did not hamper the hybridization within Sarcococca genotypes. Our findings will contribute to a more efficient breeding program and a faster achievement of hybrids with an added value.     

Sex distinction of asparagus by loop-mediated isothermal amplification and observation of seedling phenotypes

Asparagus (Asparagus officinalis L.) is a dioecious plant. In general, male and female plants are used for open-field culture and intensive cultivation, respectively. Farmers distinguish between the sexes by observing the form of the flower organs. However, because flowering begins 2?C3 years after planting, the sexes cannot be differentiated at transplantation by using this method, and planting of an all-male population is not possible. In this study, the usefulness of loop-mediated isothermal amplification (LAMP), a simple method of gene amplification, for sex distinction at the DNA level was determined. In addition, the phenotypic differences in seeds and seedlings of male and female plants were investigated for application as a method of early sex distinction. By using the LAMP method, the sex could be correctly identified in 100% of the seedlings, suggesting that this method is effective for sex distinction at the gene level. Principal component analysis was conducted with 11 selected parameters after investigating the seeds and seedlings of both male and female plants. The results revealed that male plants tend to have many stalks or cladophylls and female plants tend to have large plant forms, suggesting that the sexes can be distinguished by the external appearance of the seedlings before planting. LAMP and observation of the seedling phenotypes could be useful methods of sex distinction for increasing the efficiency of asparagus breeding.     

Bph15在非洲栽培稻、药用野生稻和宽叶野生稻中的BAC-FISH比较物理定位

用覆盖抗褐飞虱基因Bph15的两个BAC克隆，即20M14 (27 kb)和64O9 (36 kb)作为探针，对非洲栽培稻、药用野生稻和宽叶野生稻体细胞染色体进行荧光原位杂交(FISH)。两个BAC克隆均被定位于非洲栽培稻和药用野生稻第4染色体的短臂上，杂交信号的百分距离分别为37.03±4.11和81.22±3.62，相应的信号检出率为41.18%和38.22%。在宽叶野生稻中，有两对同源染色体同时检出信号，分别定位于染色体的短臂和着丝粒区，信号距着丝粒平均百分距离分别为87.78±5.23和0，信号检出率为52.58%。由此推知，这两个BAC克隆在非洲栽培稻和药用野生稻的第4染色体分布同线且同区，并且在宽叶野生稻的DD基因组也存在Bph15基因的同源序列。在未封阻的情况下，BAC克隆在非洲栽培稻的多条染色体上有杂交信号，表明它和栽培稻C_0t-1 DNA在一定程度上具有同源性。上述结果初步显示Bph15在3个稻种染色体中的相对位置。文章讨论了Bph15在3个种间的关系，为有效分离和利用Bph15基因提供了有益的依据，对不同基因组及二倍体和四倍体中功能基因可能进化机制的分析提供了线索。     

Variation in the wheat AP2 homoeologs,the genes underlying lodicule development

The bread wheat genome harbors three homoeologs of the barley gene HvAP2, which determines the cleistogamous/non-cleistogamous flowering. The three homoeologs, TaAP2-A, TaAP2-B and TaAP2-D, are derived from the A, B and D genomes. The importance of lodicule swelling in assuring non-cleistogamous flowering in a range of wild and domesticated wheat accessions of varying ploidy level was established. Re-sequencing of wheat AP2 homoeologous genes was carried out to identify natural variation at both the nucleotide and polypeptide level. The sequences of wheat AP2 homoeologs are highly conserved even across different ploidy levels and no functional variants at the key miR172 targeting site were detected. These results indicate that engineering of cleistogamous wheat will require the presence of a functional TaAP2 modification at each of the three homoeologs.     

The generation of novel species hybrids between garden dahlias and Dahlia macdougallii to increase the gene pool for variety breeding

To transfer new traits into the gene pool of garden dahlias (Dahlia variabilis), crosses between garden dahlias (2n = 64) and the epiphytic species Dahlia macdougallii (2n = 32) from the section Epiphytum were conducted. Six hybrid plants were obtained. The hybrid status was verified using three SSR markers. In addition, flow cytometry was performed to determine the genome size of the hybrids and to show that the hybrids were hexaploid as expected from crosses between tetraploids and octoploids. The open pollinated progeny of the hybrids produced four progeny with octoploid genomes. The hybrids exhibited indeterminate vegetative growth and the formation of flowers from axillary buds, similar to the father D. macdougallii. This result is of interest for breeding new varieties of dahlia with traits that are not present in the current gene pool.     

Rj (rj) genes involved in nitrogen-fixing root nodule formation in soybean

It has long been known that formation of symbiotic root nodules in soybean (Glycine max (L.) Merr.) is controlled by several host genes referred to as Rj (rj) genes, but molecular cloning of these genes has been hampered by soybean’s complicated genome structure and large genome size. Progress in molecular identification of legume genes involved in root nodule symbiosis have been mostly achieved by using two model legumes, Lotus japonicus and Medicago truncatula, that have relatively simple and small genomes and are capable of molecular transfection. However, recent development of resources for soybean molecular genetic research, such as genome sequencing, large EST databases, and high-density linkage maps, have enabled us to isolate several Rj genes. This progress has been achieved in connection with systematic utilization of the information obtained from molecular genetics of the model legumes. In this review, we summarize the current status of knowledge of host-controlled nodulation in soybean based on information from recent studies on Rj genes, and discuss the future research prospects.     

In silico-selection of Brassica rapa organelle genome-derived BACs using their end sequences and sequence level comparative analysis of the 124 kb mitochondrial genome sequences in the family Brassicaceae

We identified BAC clones which harbor DNAs derived from the B. rapa organelle genomes by in silico mapping of 80,292 B. rapa BAC end sequences on the Arabidopsis organelle genomes and subsequent insert size estimation and fingerprinting. A total of 1,048 putative chloroplast genome-derived BAC clones (2.6%) were identified. Fingerprinting and sequencing revealed that many of them represented the entire chloroplast genome (about 150 kb). Meanwhile, only 59 putative mitochondrial genome-derived BACs (0.15%) were identified and most of them showed rare agreement between the in silico map and fingerprinting. We sequenced BAC clone KBrB042G11 (42G11) and compared it to the mitochondrial genome of B. napus and A. thaliana which showed dynamic rearrangement events. The order of 33 orthologous genes was collinear between the 42G11 BAC and its counterpart in B. napus. Five distinctive rearrangements and two InDels were identified between these two closely related species and the rearrangements were related to the occurrence of small tandem repeat sequences. Sequences of the 33 orthologous genes in the homoeologous regions of B. napus and B. rapa were almost 100% identical. Gene orders showed no colinearity between Arabidopsis and Brassica even though 31 orthologous genes shared high sequence similarity with p-values over 1E-32. FISH analysis using the identified BAC revealed a large chloroplast genome insertion in the pericentromeric region of chromosome (chr.) 4 of B. rapa.     

Genome-specific markers of tetraploid wheats and their putative diploid progenitor species

The objective of this study was to isolate genome‐specific markers from the genomes of tetraploid wheats and the putative donor diploid species on the basis of random amplified polymorphic DNA analysis followed by cross‐hybridization. Twenty different Triticum and Aegilops species and accessions were analysed by polymerase chain reaction (PCR) using 30 random primers. The polymorphic PCR fragments were then isolated, labelled and used in cross‐hybridization screenings. The hybridization results established that one marker was specific to the Ae. speltoides S genome, two to the A genome, one to the B genome and five to the G genomes of polyploid species (and to the genomes of the corresponding progenitor species). Four markers were identified that were specific to both the B and G genomes. Analysis of the Triticum and Aegilops species and accessions supported the notion that Ae. speltoides is more closely related to the B and G genomes of polyploid wheat species than were other members of the Sitopsis section. The data also indicated that the B and G genomes had originated from different accessions of Ae. speltoides.     

Variation in floral scent compounds recognized by honeybees in Brassicaceae crop species

Floral scent attracts pollinators. We investigated the floral scent compounds recognized by pollinators in six Brassica crop species, including allogamous species with different genomes and autogamous species with two parental genomes and radish (Raphanus sativus). Biologically active compounds recognized by honeybees were screened from all floral compounds by combined gas chromatography–electroantennogram analysis and their profiles were determined by gas chromatography–mass spectrometry. Fourteen of the 52 compounds were active. All accessions had more than two active compounds, but the compounds greatly differed between the two genera. On the basis of similarities in whether active compounds were presence or absence, their amount and their composition ratio, we divided the Brassica accessions into three to five groups by cluster analyses. Most groups were composed of a mixture of allogamous and autogamous species sharing same genome, indicating that the variation depended on genome, not species. These results suggest that all species require pollinator visits for reproduction, despite their different reproductive systems. However, the inter-genus and intra-specific variations shown by the multiple groups within a species might cause different visitation frequencies by pollinators between genera and among accessions within a species, resulting in insufficient seed production in some accessions or species.     

Sex chromosomes and quantitative sex expression in monoecious hemp (Cannabis sativa L.)

Hemp (Cannabis sativa) has a highly variable sexual phenotype. In dioecious hemp, the sex is controlled by heteromorphic sex chromosomes according to an X-to-autosomes equilibrium. However, in monoecious hemp, the sex determinism remains widely unknown and has never been related to a quantitative approach of sex expression. The present paper aims to contribute to the comprehension of the sex determinism in monoecious hemp by assessing the genotypic variability of its sex expression and establishing its sex chromosomes. Five monoecious and one dioecious cultivars were grown in controlled conditions under several photoperiods. The monoecy degree of 194 monoecious plants was recorded at each node by a figure ranging from 0 (male flowers only) to 6 (female flowers only). The genome size of 55 plants was determined by flow cytometry. The DNA of 115 monoecious plants was screened with the male-associated marker MADC2. The monoecy degree varied significantly among monoecious cultivars from 3.36 ± 2.28 in ‘Uso 31’ to 5.70 ± 0.81 in the most feminised ‘Epsilon 68’. The variation of monoecy degree among cultivars remained consistent across trials despite a significant “cultivar × trial” interaction and partly agreed with their earliness. The genome size of monoecious plants (1.791 ± 0.017 pg) was not different from that of females (1.789 ± 0.019 pg) but significantly lower than that of males (1.835 ± 0.019 pg). MADC2 was absent from all monoecious plants. These results strongly support that cultivars of monoecious hemp have the XX constitution and that their sex expression has a genetic basis.