Utilization Of Germplasm Resources Research Articles

Establishment and population genetic analysis of SNP fingerprinting of Chinese soft-shelled turtle (Pelodiscus sinensis)

Chinese soft-shelled turtle (Pelodiscus sinensis) is an important aquaculture species in China. However, with the continuous development of P. sinensis cultures, problems related to germplasm degradation and population mixing have become increasingly prominent. This has seriously affected the protection and utilization of germplasm resources by P. sinensis. To broaden the understanding of the germplasm resources of P. sinensis and improve the efficiency of germplasm identification, Super-GBS sequencing technology was used to conduct population genetic analysis of P. sinensis samples from six different populations distributed in five provinces. As a result, 1464,488 SNPs were identified. Genetic structure analysis revealed that the six P. sinensis populations could be divided into three sub-populations. The genetic differentiation coefficient of the DT and CY populations was very low (Fst=0.0088), indicating that differentiation had not yet occurred. Additionally, 15 KASP core markers for the identification of P. sinensis populations were developed using KASP typing technique for the first time. Genetic information can lay a foundation for the protection and utilization of germplasm resources by P. sinensis, and the fingerprinting database developed in this study provides a practical tool for authenticating P. sinensis.

Core collection construction of tea plant germplasm in Anhui Province based on genetic diversity analysis using simple sequence repeat markers

The tea plant [Camellia sinensis (L.) O. Kuntze] is an industrial crop in China. The Anhui Province has a long history of tea cultivation and has a large resource of tea germplasm with abundant genetic diversity. To reduce the cost of conservation and utilization of germplasm resources, a core collection needs to be constructed. To this end, 573 representative tea accessions were collected from six major tea-producing areas in Anhui Province. Based on 60 pairs of simple sequence repeat (SSR) markers, phylogenetic relationships, population structure and principal coordinate analysis (PCoA) were conducted. Phylogenetic analysis indicated that the 573 tea individuals clustered into five groups were related to geographical location and were consistent with the results of the PCoA. Finally, we constructed a core collection consisting of 115 tea individuals, accounting for 20% of the whole collection. The 115 core collections were considered to have a 90.9% retention rate for the observed number of alleles (Na), and Shannon’s information index (I) of the core and whole collections were highly consistent. Of these, 39 individuals were preserved in the Huangshan area, accounting for 33.9% of the core collection, while only 10 individuals were reserved in the Jinzhai County, accounting for 8.9% of the core set. PCoA of the accessions in the tea plant core collection exhibited a pattern nearly identical to that of the accessions in the entire collection, further supporting the broad representation of the core germplasm in Anhui Province. The results demonstrated that the core collection could represent the genetic diversity of the original collection. Our present work is valuable for the high-efficiency conservation and utilization of tea plant germplasms in Anhui Province.

Morphological to Molecular Markers: Plant Genetic Diversity Studies in Walnut (Juglans regia L.)—A Review

Persian walnut populations have tremendous morphological and allelic diversity in their germplasm due to heavy outcrossing and years of seed multiplication. These variations are assessed by morphological, cytological, biochemical, and molecular markers. Various researchers have used different tree, foliage, flower, nut, and kernel traits to evaluate morphological/phenotypic diversity. In walnut, morphological indices are first taken into consideration to describe and classify the germplasm, but the environment influences them. In comparison, DNA-based markers can detect genetic diversity at any stage of plant development and have been shown to be a potential tool for assessing variation at the DNA level and deciphering genetic relationships within and between species. Microsatellites are very powerful and informative among DNA-based markers in studying genetic relationships and genetic identity at different levels. They are neutral, highly frequent, uniformly distributed, hypervariable, codominant, highly reproducible, produce many alleles per locus, and require a small amount of DNA for analysis. Current breeding objectives can be achieved by selecting superior genotypes from the germplasm, supplemented by molecular characterization in the selection of parents for each breeding program. Therefore, the use of morphological and molecular markers is recommended for efficient exploration and utilization of germplasm resources and to improve diversity among genetic resources. The published literature on morphological and molecular markers, especially simple sequence repeats (SSRs), is presented in this review to provide current insights into the level of genetic diversity in walnut.

Developments on Core Collections of Plant Genetic Resources: Do We Know Enough?

The core collection is a small subset that minimizes genetic redundancy while preserving the maximum genetic diversity of the entire population. Research on the core collection is crucial for the efficient management and utilization of germplasm resources. In this paper, the concept of the core collection and the research progress of its construction have briefly been summarized. Subsequently, some perspectives have been proposed in this research field for the near future. Four novel opinions have been presented, (1) the effective integration of multiple data types and accurate phenotyping methods need to be focused on; (2) the sampling strategy and bioinformatics software should be given attention; (3) the core collection of afforestation tree and bamboo species, with a wide natural distribution range and a large planting area, need to be carried out as soon as possible; (4) we should place a high priority on the study of genes discoveries and utilize these with a rapid, precise and high-throughput pattern based on re-sequencing technology. This paper provides a theoretical and technological reference for further study and the application of the plant core collection.

Evaluating the genetic diversity of Erythropalum scandens based on using inter-simple sequence repeat markers

Erythropalum scandens Blume, an emerging medicinal plant with great potential for drug development, also possesses high edible value. In this study, we investigated the genetic diversity of the germplasm of E. scandens obtained from different geographical locations using inter simple sequence repeat (ISSR) markers. For this purpose, 18 ISSR primer pairs with a distinct background and adequate polymorphism were selected. We established an optimal ISSR–PCR reaction system (20 µL) with the following parameters: 1 µL DNA template (60 ng µL−1), 1.2 µL primers (10 µmol µL−1), 10 µL MasterMix, and 7.8 µL H2O. A total of 183 loci were amplified using the 18 primer pairs, of which 121 (66.12%) indicated polymorphism. Moreover, 34 germplasms of E. scandens exhibited genetic similarity coefficients ranging from 0.7104 to 0.9563, genetic distances ranging from 0.0447 to 0.3420, Nei’s genetic diversity index of 0.1946, and Shannon’s information index of 0.2982, suggesting high intraspecific genetic diversity. UPGMA cluster and PCoA analyses distinguished the germplasm of E. scandens obtained from Guangxi from those collected from Guangdong, Hainan, Fujian, and Guizhou. However, the Mantel correlation analysis revealed that the genetic variation among the 34 germplasms of E. scandens was not significantly related to geographical distance. The analysis of the genetic background of wild and cultivated germplasms of E. scandens can help guide variety selection and breeding. Furthermore, the present study revealed the genetic background and affinities among 34 germplasms of E. scandens. Overall, our findings lay the foundation for the conservation and utilization of germplasm resources, identification and classification of varieties, and variety selection and improvement of E. scandens at the molecular level.

Studies on Pollen Morphology, Pollen Vitality and Preservation Methods of Gleditsia sinensis Lam. (Fabaceae)

Gleditsia sinensis Lam. (Fabaceae) is an endemic species in China, which has a wide range of ecological functions and high economic value. G. sinensis belongs to androdioecy, and the stamens of perfect flowers are aborted, meaning that a perfect flower is a functional female flower. Understanding the dynamic process of flowering and the characteristics of pollen morphology effectively determine the viability of pollen vitality, and the suitable conditions for short-term storage of pollen can provide theoretical basis and technical reference for hybrid breeding and germplasm conservation of G. sinensis. In this study, the male plants of G. sinensis in Guiyang area were used as research materials. The flowering dynamic process of male flowers was recorded through field observation. The morphology of pollen was observed and analyzed with a scanning electron microscope (SEM). The germination characteristics of pollen were studied with an in vitro germination method, and the pollen vitality was also determined using four staining methods. The effects of different storage temperatures and water contents on pollen germination rate were discussed. The results showed that the male flowers of G. sinensis had a short, single flowering period, lasting 2–3 days from the opening to the shedding. The dynamic opening process of a single flower was artificially divided into five stages. Pollen grains of G. sinensis are oblate spheroidal, tricolporate with equatorial elongated endoapertures and the sporoderm surface is reticulate. The MTT (Thiazolyl Blue Tetrazolium Bromide) staining method could accurately and quickly determine the pollen vitality of G. sinensis. The highest pollen germination rate was 65.89% ± 3.41%, and the length of the pollen tube was 3.96 mm after cultured in 15% sucrose + 100 mg/L boric acid + 20 mg/L calcium chloride for 24 h. It was necessary to collect the pollen at the big bud stage, which was conducive to improving the efficiency of pollen collection because the pollen had been mature with high pollen vitality at this stage. When it came to pollen preservation, the pollen germination rate was significantly affected by storage time, storage temperature and pollen water content. The pollen still had high vitality after being stored at −80 °C for 30 days when the moisture content of the pollen decreased to 9%, and the pollen germination rate only decreased by 28.84% compared with that before storage. In conclusion, this study has comprehensively and systematically studied the morphology, vitality determination and preservation methods of the pollen of G. sinensis, providing a theoretical basis for the cross regional breeding and the conservation and utilization of germplasm resources.

Study on the Protection System and Capacity Building of Crop Germplasm Resources

Although remarkable achievements have been made in the protection and utilization of agricultural germplasm resources over the years, there are still problems. This paper summarizes the current situation of the protection and utilization of crop germplasm resources in Yunnan Province and puts forward development ideas and suggestions for the protection and utilization of resources in the future. It is pointed out that the protection and utilization of germplasm resources are not systematic, the utilization rate of germplasm resources is low, breeding innovation is difficult, there is a lack of large-scale "breeding and promotion integration" enterprises, the infrastructure is not perfect, and the use of land for supporting facilities is difficult, and relevant suggestions are put forward in order to provide reference for the protection and utilization of germplasm resources, the selection and promotion of excellent varieties and the development of seed industry in China.

Current Situation and Utilization of Velvet Deer Germplasm Resources in China.

Velvet deer are not only a representative special economic animal but also an important part of livestock. With the increasing awareness of international competition for germplasm resources in China, more and more attention has been paid to the protection and utilization of germplasm resources. However, there is poor understanding about velvet deer resources. Therefore, we are providing a comprehensive introduction of Chinese velvet deer germplasm resources from the aspects of ecological distribution, domestication and breeding.

Chloroplast genome in Malus floribunda Siebold.

Malus floribunda Siebold. (Malus) is widely cultivated all over the world, which is of high ornamental value and breeding significance. Comparative analysis of the chloroplast genome can help enrich the phylogenetic relationship and facilitate germplasm utilization of Malus. Based on the whole genome sequencing data, a complete chloroplast genome (M. floribunda) with tetrad structure was assembled. The chloroplast genome (160 037 bp) was composed of a large single-copy (LSC) region (88 142 bp), inverted repeat (IR) B (26 353 bp), a small single-copy (SSC) region (19 189 bp), and IRA (26 353 bp). A total of 111 genes were annotated: 78 protein-coding genes, 29 tRNA genes and 4 rRNA genes. In addition, a large number of repeat sequences were identified in the genome, which was slightly different from that of M. sieboldii and M. toringoides. As for the relative synonymous codon usage, 30 high-frequency codons were found, and the codons tended to end with A/T. The results of interspecific sequence alignment and boundary analysis suggested the sequence variation of the LSC region was large, and the expansion and contraction of the SC region and IR region of the eight Malus species were generally similar. According to the phylogenetic analysis of chloroplast genome sequences, M. floribunda, M. hupehensis, and M. toringoides were grouped into one clade. The findings in this study can provide data support for the development of genetic markers and utilization of germplasm resources in the future.

Population Genetic Diversity and Differentiation of Mitten Crab, Genus Eriocheir, Based on Microsatellite Markers

Mitten crab, Eriocheir sensu stricto, is an important indigenous aquatic species and food source in Eastern Asia. Genetic diversity is an important prerequisite for the conservation and utilization of germplasm resources. However, the genetic diversity and differentiation of Eriocheir s. s. remains unclear. This study evaluated population genetic diversity and genetic differentiation of three Eriocheir s. s. populations from Suifenhe (SFH), Liaohe (LH), and Nanliujiang (NLJ), China, based on 19 microsatellite markers. The mean observed number of alleles (Na) was 22.84 alleles, and the mean polymorphism information content (PIC) was 0.86, which demonstrated high genetic diversity. The allele frequency distribution showed an “L” shape, ranging from 0.01 to 0.74. The genetic diversity parameter values of the LH population were higher than those of the other two populations. All pairwise FST values showed significant differences among the three Eriocheir s. s. populations (p < 0.01). The value of Nei’s genetic distance (DS) varied from 0.31 (between SFH and NLJ populations) to 0.33 (between SFH and LH populations). Overall, The results illustrate that the three wild Eriocheir s. s. populations have high genetic diversity and high genetic differentiation. This study provides the basis for Eriocheir s. s. utilization in China.

Complete chloroplast genome sequence of Kosteletzkya pentacarpos

Kosteletzkya pentacarpos is a promising plant being developed as a salt-tolerant biofuel crop that also the ability to fix heavy metals. Here, high-throughput sequencing technology was used to sequence and assemble the chloroplast genome of K. pentacarpos. The full length of the chloroplast genome is 161,777 bp, comprising a large single-copy region of 90,019 bp, a small single-copy region of 18,978 bp, and a pair of inverted repeats of 26,390 bp. A total of 113 genes were annotated, including 79 protein-coding, 30 transfer RNA, and 4 ribosomal RNA genes. Phylogenetic analysis based on whole chloroplast genome sequences showed that K. pentacarpos has a close relationship with Abelmoschus in Malvaceae. This study increases the available genomic information on K. pentacarpos, and provides a basis for the rational exploitation and utilization of germplasm resources.

The complete mitochondrial genome of a vulnerable mandarin fish Coreoperca liui (Teleostei: Perciformes: Serranidae) from Qiandaohu Lake in China

Coreoperca liui as an approximate species of the genus Siniperca, provides an important source for the genetic diversity of the mandarin fish, which is valuable for the protection of biodiversity and utilization of germplasm resources. The complete mitochondrial genome of C. liui is 16,482 bp long and it consists of 13 protein-coding genes (PCGs), two ribosomal RNA genes, 22 transfer RNA genes, and a control region (D-loop). Phylogenetic analysis using the maximum-likelihood method, based on 13 PCGs and two rRNA from 13 species produced three major clades. The phylogenetic tree showed that C. liui is most closely related to Coreoperca whiteheadi. Our results provide useful information for understanding the phylogeny of the genus Coreoperca, as well as for conducing conservation studies of Sinipercidae and related species.

Development of a Model for Genomic Prediction of Multiple Traits in Common Bean Germplasm, Based on Population Structure

Due to insufficient identification and in-depth investigation of existing common bean germplasm resources, it is difficult for breeders to utilize these valuable genetic resources. This situation limits the breeding and industrial development of the common bean (Phaseolus vulgaris L.) in China. Genomic prediction (GP) is a breeding method that uses whole-genome molecular markers to calculate the genomic estimated breeding value (GEBV) of candidate materials and select breeding materials. This study aimed to use genomic prediction to evaluate 15 traits in a collection of 628 common bean lines (including 484 landraces and 144 breeding lines) to determine a common bean GP model. The GP model constructed by landraces showed a moderate to high predictive ability (ranging from 0.59–0.88). Using all landraces as a training set, the predictive ability of the GP model for most traits was higher than that using the landraces from each of two subgene pools, respectively. Randomly selecting breeding lines as additional training sets together with landrace training sets to predict the remaining breeding lines resulted in a higher predictive ability based on principal components analysis. This study constructed a widely applicable GP model of the common bean based on the population structure, and encouraged the development of GP models to quickly aggregate excellent traits and accelerate utilization of germplasm resources.

Characterizing Tetraploid Populations of Actinidia chinensis for Kiwifruit Genetic Improvement.

Understanding genetic diversity and structure in natural populations and their suitable habitat response to environmental changes is critical for the protection and utilization of germplasm resources. We evaluated the genetic diversity and structure of 24 A. chinensis populations using simple sequence repeat (SSR) molecular markers. The potential suitable distribution of tetraploid A. chinensis estimated under the current climate and predicted for the future climate was generated with ecological niche modeling (ENM). The results indicated that the polyploid populations of A. chinensis have high levels of genetic diversity and that there are distinct eastern and western genetic clusters. The population structure of A. chinensis can be explained by an isolation-by-distance model. The results also revealed that potentially suitable areas of tetraploids will likely be gradually lost and the habitat will likely be increasingly fragmented in the future. This study provides an extensive overview of tetraploid A. chinensis across its distribution range, contributing to a better understanding of its germplasm resources. These results can also provide the scientific basis for the protection and sustainable utilization of kiwifruit wild resources.

Creation and validation of core subset of potato (Solanum tuberosum) germplasm

Potato (Solanum tuberosum L.) is the only non-cereal staple food crop globally. The progress of this temperate crop in the sub-tropical country relies strongly on improvement activities utilizing the genetic resources acquired over a time period. However, proper maintenance, evaluation and utilization of germplasm resources is a cumbersome task. Creation of core set of conserved genetic resources is an efficient and cost effective method of management of conserved germplasm and its use in breeding program. Indian collection of potato (Solanum tuberosum ssp tuberosum) constitutes 1399 accessions emanating from the five continents maintained under ex situ conservation. A core subset of 140 potato germplasm was developed using Power Core software based on 19 morphological traits. Majority of entries in the core subset were from South America (42.14%). Comparison of mean data using Newman-Keul's test and variances using Levene's test for different traits revealed representation of actual diversity of base population by the core subset. Correlations among the traits governed by co-adapted gene complexes were conserved in the core subset. Shannon-Weaver diversity indices indicate that the potato core subset maximized the phenotypic diversity of the potato germplasm. Validation of core subset using principal component analysis reveals that the first 4 PCs represents 55.27% of total variations in core subset compared to 52.50% that in entire collection. The identified core subset can be used in Indian potato improvement programs as well as in genomics studies.

The physiology of plant seed aging: a review

Seed quality plays an important role in the agricultural and animal husbandry production, the effective utilization of genetic resources, the conservation of biodiversity and the restoration and reconstruction of plant communities. Seed aging is a common physiological phenomenon during storage. It is a natural irreversible process that occurs and develops along with the extension of seed storage time. It is not only related to the growth, yield and quality of seed and seedling establishment, but also has an important effect on the conservation, utilization and development of plant germplasm resources. The physiological mechanisms of seed aging are complex and diverse. Most studies focus on conventional physiological characterization, while systematic and comprehensive in-depth studies are lacking. Here we review the recent advances in understanding the physiology of seed aging process, including the methods of seed aging, the effect of aging on seed germination, and the physiological and molecular mechanisms of seed aging. The change of multiple physiological parameters, including seed vigor, electrical conductivity, malondialdehyde content and storage material in the seed, antioxidant enzyme activity and mitochondrial structure, were summarized. Moreover, insights into the mechanism of seed aging from the aspects of transcriptome, proteome and aging related gene function were summarized. This study may facilitate the research of seed biology and the conservation and utilization of germplasm resources.

Analysis of genetic diversity in Prunus sibirica L. in inner Mongolia using SCoT molecular markers

Population genetic diversity contributes to the protection and utilization of germplasm resources, especially via genetic breeding. In the present study, start codon targeted polymorphism (SCoT) molecular markers were used to study the genetic diversity of 278 individuals from 10 Prunus sibirica L. populations in Inner Mongolia. A total of 289 polymorphic bands were amplified with 23 SCoT primers, showing a polymorphism percentage of 98.87% and an average of 12.6 polymorphic bands per primer. The SCoT21, SCoT32, and SCoT53 primers amplified up to 17 bands, and the polymorphism percentage was 100%. The minimum number of bands amplified by SCoT25 was 9, and the polymorphism percentage was 90%. Therefore, SCoT molecular markers were shown to be highly polymorphic and suitable for genetic diversity studies of P. sibirica in Inner Mongolia. The analysis of molecular variance showed that 39% of the observed genetic differentiation occurred among populations and 61% occurred within populations, indicating that the genetic differentiation within populations was greater than that among populations. The results of the unweighted pair-group method with an arithmetic cluster analysis, principal coordinate analysis and STRUCTURE analysis were basically the same and divided the 278 individuals from the 10 populations into 2 groups. The results indicated that the efficient SCoT molecular marker-based genetic diversity analysis of P. sibirica in Inner Mongolia can provide a reference for P. sibirica variety breeding and resource development.

Development and characterization of 37 SNP markers for the largemouth bass (Micropterus salmoides) by using PCR–RFLP method

Largemouth bass (Micropterus salmoides) is an economically important species in China. Contrary to its rapidly increasing yield during the last decades, the domestic genetic diversity of largemouth bass has gradually declined. In this study, 37 single nucleotide polymorphism (SNP) markers were developed based on genotyping-by-sequencing data and characterized by genotyping 32 individuals using the PCR–restriction fragment length polymorphism method. The effective number of alleles (Ne), observed heterozygosity (Ho), expected heterozygosity (He), and polymorphic information content (PIC) of these SNPs ranged from 1.168 to 1.998, 0.156 to 0.844, 0.146 to 0.507, and 0.134 to 0.375, respectively. Totally, five loci deviated significantly from Hardy–Weinberg equilibrium (p < 0.05), while there existed no linkage disequilibrium at all loci. These novel polymorphic markers will lay the foundation for future rationally excavation and utilization of germplasm resources in M. salmoides.

Strategy for the construction of a core collection for Pinus yunnanensis Franch. to optimize timber based on combined phenotype and molecular marker data

The core collection construction strategies are critical for the conservation and utilization of germplasm resources. The combined data of phenotypic characteristics and SRAP markers in 780 samples were employed to construct a core collection for Pinus yunnanensis Franch. Based on geographical distribution, a total of four P. yunnanensis germplasm subsets were configured by improving a least distance stepwise sampling method at four sampling proportions. The representativeness of germplasm subsets for original germplasms were evaluated by quantitative and qualitative trait independence tests. For the quantitative traits test, the germplasm subset at a 30% sampling proportion was the best of four germplasm subsets by the mean t-test, the variance F-test, the frequency distribution χ2-test, the Shannon diversity index, and five effective evaluation parameters. For the qualitative traits test, there were significant differences between the original germplasm and germplasm subset at a 10% sampling proportion in the mean t-test of genetic diversity indicators. The genetic diversity within the population was significantly different between the original germplasm and germplasm subsets at 20% and 10% sampling proportions, respectively, in the variance F-test. The results of quantitative and qualitative traits tests revealed that the 30% sampling proportion was appropriate for the construction of the core collection.

Genetic diversity analysis in Chinese miniature pigs using swine leukocyte antigen complex microsatellites.

ObjectiveThe swine leukocyte antigen (SLA) gene group, which is closely linked and highly polymorphic, has important biomedical significance in the protection and utilization of germplasm resources. However, genetic polymorphism analyses of SLA microsatellite markers in Chinese miniature pigs are limited.MethodsEighteen pairs of microsatellite primers were used to amplify the SLA regions of seven miniature pig breeds and three wild boar breeds (n = 346) from different regions of China. The indexes of genetic polymorphism, including expected heterozygosity (He), polymorphic information content (PIC), and haplotype, were analyzed. The genetic differentiation coefficient (Fst) and neighbor-joining methods were used for cluster analysis of the breeds.ResultsIn miniature pigs, the SLA I region had the highest numbers of polymorphisms, followed by the SLA II and SLA III regions; the region near the centromere had the lowest number of polymorphisms. Among the seven miniature pig breeds, Diannan small-ear pigs had the highest genetic diversity (PIC value = 0.6396), whereas the genetic diversity of the Hebao pig was the lowest (PIC value = 0.4330). The Fst values in the Mingguang small-ear, Diannan small-ear, and Yunnan wild boars were less than 0.05. According to phylogenetic cluster analysis, the South-China-type miniature pigs clustered into one group, among which Mingguang small-ear pigs clustered with Diannan small-ear pigs. Haplotype analysis revealed that the SLA I, II, and III regions could be constructed into 13, 7, and 11 common haplotypes, respectively.ConclusionThis study validates the high genetic diversity of the Chinese miniature pig. Mingguang small-ear pigs have close kinship with Diannan small-ear pigs, implying that they may have similar genetic backgrounds and originate from the same population. This study also provides a foundation for genetic breeding, genetic resource protection, and classification of Chinese miniature pigs.