Analysis of Equivalence Between Traditional and Nontraditional Medicinal Varieties of Citri Reticulatae Semen from Sichuan Province / 中国实验方剂学杂志

Objective:To study the differences in genetic relationship， shape， size， and flavonoid content between traditional and nontraditional medicinal varieties of Citri Reticulatae Semen produced in Sichuan province as well as their equivalence. Method:Six batches of traditional medicinal Citri Reticulatae Semen （<italic>Citrus reticulata</italic> 'Dahongpao'） and 23 batches of nontraditional medicinal varieties were collected， and their genetic relationship was explored using sequence-related amplified polymorphism （SRAP） markers. Following the observation of their shapes and sizes under a stereomicroscope， the contents of naringin， hesperidin， and neohesperidin were measured by high performance liquid chromatography （HPLC）. SIMCA 14.1 software was used for cluster analysis of their shapes， sizes， and flavonoid contents， thus figuring out the similarities between the traditional and nontraditional medicinal varieties in character， size， and chemical components. Result:SRAP markers-based genetic relationship analysis effectively distinguished different Citri Reticulatae Semen varieties from each other. Some samples collected from the same or adjacent places exhibited a close genetic relationship and they shared high similarities in shape， size， and flavonoid content. However， the traditional medicinal Citri Reticulatae Semen was still quite different from most nontraditional medicinal varieties. Conclusion:The analysis of differences in genetic materials， appearance， character， and active ingredient content between the traditional and nontraditional medicinal varieties revealed that the equivalence<italic> </italic>of <italic>C.</italic> <italic>reticulata</italic> 'Ponkan' samples from some regions with the traditional medicinal variety was the largest， enabling them to be considered as the emerging medicinal variety.

Optimization of SRAP-PCR System for Valeriana officinalis var. latifolia and Primer Screening / 中国实验方剂学杂志

Objective:To establish the sequence-related amplified polymorphism （SRAP）-polymerase chain reaction （PCR） system for <italic>Valeriana officinalis</italic> var. <italic>latifolia</italic>，so as to lay the theoretical and technical foundations for the breeding of<italic> V. officinalis </italic>var. <italic>latifolia</italic>. Method:Single factor test was applied to investigate the effects of <italic>Taq</italic> Mix dose，Mg²⁺concentration，template DNA concentration，and <italic>Taq </italic>DNA polymerase content on SRAP-PCR amplification of <italic>V. officinalis </italic>var. <italic>latifolia</italic>，based on which the orthogonal experiments were performed to optimize the SRAP-PCR system for <italic>V. officinalis </italic>var. <italic>latifolia</italic>. The effective primers that could be used for genetic diversity studies of <italic>V. officinalis</italic> var. <italic>latifolia </italic>were selected under the optimal reaction condition. Result:The results of the single factor test showed that <italic>Taq </italic>Mix dose within the range of 8-11 μL resulted in better amplification. The addition of a low concentration of Mg²⁺，the medium to low concentrations of template DNA，or the low concentration of <italic>Taq</italic> DNA polymerase enhanced the amplification efficiency or richness. As demonstrated by the orthogonal experiments，the influencing degrees of related factors on SRAP-PCR amplification of <italic>V. officinalis</italic> var. <italic>latifolia </italic>were sorted in a descending order as follows： <italic>Taq</italic> Mix dose><italic>Taq</italic> DNA polymerase content>Mg²⁺ concentration>template DNA concentration. The optimal reaction system for <italic>V. officinalis</italic> var. <italic>latifolia </italic>was determined to consist of 11 μL of <italic>Taq</italic> Mix，30 ng of template DNA，0.025 mmol·L^-1Mg²⁺，1.5 U<italic> </italic>of<italic> Taq </italic>DNA polymerase，5 μmol·L^-1 forward primer，and 5 μmol·L^-1 reverse primer，which was supplemented to 20 μL with ddH₂O. The optimal annealing temperature was 36.8 ℃. A total of 17 pairs of effective primers with high band resolution and polymorphism were selected from 88 primer pairs for SRAP-PCR of <italic>V. officinalis</italic> var. <italic>latifolia</italic>. Conclusion:The established SRAP-PCR system for <italic>V. officinalis</italic> var. <italic>latifolia</italic> is stable， which can be used for genetic diversity studies of <italic>V. officinalis</italic> var. <italic>latifolia</italic>.

Establishment of DNA fingerprints for Chrysosplenium using SRAP Markers / 中国中药杂志

As a traditional Chinese medicinal material, Chrysosplenium is urgently needed for genetic resource investigation and protection research due to the decrease of its wild resources in recent years. After investigating the wild resources, we conducted genetic polymorphism and clustering studies of 24 species(a total of 36 samples) of Chrysosplenium using SRAP technique. The results showed that a total of 374 polymorphic bands were obtained using 18 pairs of SRAP primers to amplify these samples, on average of 20.7 bands for each primer pair. We used the biological software to analyze the population's genetic parameter and got the N_a value as 2.000 0, N_(e )value as 1.408 4, the average Nei's index as 0.263 5, and the average Shannon information index as 0.419 1. UPGMA cluster analysis showed that all the samples can be divided into three major groups at the genetic similarity coefficient of 0.70: there are 18 species(24 samples) gathered for the Ⅰ groups, 3 species or variation(7 samples) for Ⅱ groups, and 3 species(5 samples) for Ⅲ groups. The differences of these Chrysosplenium species at the molecular level are consistent with that of their geographical and ecological distribution. At the same time, we used SRAP technology to construct 36 DNA digital fingerprints of Chrysosplenium and obtained the unique molecular identification band type of each material. These results will provide effective methods and reliable basis for the identification, protection and genetic diversity analysis of the germplasm resources of Chrysosplenium, and lay a foundation for the further development and utilization of them.

Genetic diversity analysis of Gardenia jasminoides based on ISSR and SRAP molecular markers / 中草药

Objective To analyze of genetic diversity of Gardenia jasminoides and provide the information for the conservation and new variety breeding of G. jasminoides. Methods 12 ISSR primers and 9 primer combinations of SRAP were used to assess the polymorphisms, genetic diversity, and cluster analysis within 21 G. jasminoides materials from three populations. Results The results showed that 100 (80.00%) of 125 and 74 (92.50%) of 80 bands were polymorphic by ISSR and SRAP primers amplification, respectively. In ISSR results, the populations of species level of observed number of alleles (Na), effective number of alleles (Ne), Nei’s gene diversity (H), Shannon’s information index (I), total genetic diversity for species (Ht), and the mean heterozygosity with populations (Hs) were 1.461 3, 1.307 7, 0.173 1, 0.254 5, 0.239 1, and 0.173 1, respectively. Comparatively, for SRAP primers, the Na, Ne, H, I, Ht, and Hs value was 1.579 2, 1.342 1, 0.197 4, 0.295 9, 0.289 9, and 0.197 4, respectively. The coefficient of gene differentiation (Gst) for population was 0.276 2 and 0.318 9, which indicated that the within-population component accounted for 73.38% and 68.11%, respectively. The average mean of gene flow (Nm > 1) suggested that there certainly gene flow among the populations. UPGMA analysis showed that 21 samples were clustered into 2 branches, and a hierarchical dendrogram based on SRAP was more consistent with actual populations. Conclusion The gene diversity of G. jasminoides populations was high. The characteristics of genetic structure included genetic differentiation that occurs mainly within populations, which provided a reference for conservation and breeding of G. jasminoides germplasm.

Genetic diversity of endophytic fungi strains Fusarium proliferatum isolated from Belamcanda chinensis using ISSR and SRAP markers / 中草药

Objective: In order to understand the genetic diversity of endophytic fungi strain Fusarium proliferatum isolated from Belamcanda chinensis. Methods: A total of 52 ISSR primers and 90 SRAP primers were used to detect the genetic diversity among 17 F. proliferatum strains. Results: The results indicated that 27 ISSR primers and 38 SRAP primers were screened out for the genetic diversity analysis. 178 bands were amplified from 27 ISSR primers, among which 131 (63%) allelic variations were detected. However, 357 bands were amplified by 38 SRAP primers, among which 323 (91%) allelic variations were detected. The value of allelic polymorphism information content (PIC) of ISSR primers ranged from 0.19 to 0.91, with the average of 0.70 per primer. The value of PIC of SRAP primers ranged from 0.00 to 0.93, with the average of 0.72 per primer. The value of Nei's genetic similarity (GS) indexes of 17 strains based on ISSR, SRAP and ISSR + SRAP genetic locus varied from 0.73-0.99, 0.72-0.95 and 0.73-0.95, and with the average of 0.84, 0.85 and 0.85, separately. Cluster analysis showed that the 17 strains in this study could be clustered into three groups, three strains from the roots were clustered together, and F. proliferatum strains isolated from stems and leaves were gathered in other two groups. Cluster analysis revealed that genetic similarity of 17 strains were high, this suggested that the 17 strains had a near relationship, in accordance with the traditional morphology identification. Conclusion: The results show that the ISSR and SRAP technology is more efficient than traditional morphology identification. It is also found that ISSR and SRAP markers could more really reflect the genetic diversity of endophytic fungi strain F. proliferatum from B. chinensis, which can provide the basis for the application of molecular biotechnology in endophytic fungi of F. proliferatum from B. chinensis.

Investigation of Differential Genes Expression in the Genome of Epipactis flava Seidenf. (Orchidaceae) under Flooded Condition Using cDNA-SRAP Analysis

Epipactis flava Seidenf. is classified into a rare type of orchid species, a rheophyte. The specific habitat for its life cycle requires a strong current and flooding time. In this study, differential gene expression between submerged and non-submerged conditions was studied using the cDNA-SRAP method. RNAs were extracted from rhizomes at 0, 6, 12, 24 and 48 hours after submergence. Seven candidate genes were selected to study the relative gene expression by real-time RT-PCR and found to be upregulated in submergence condition. Six of these, RNA polymerase sigma factor gene, ATPase gene, Plant homeodomain finger Alfin-like genes, DNA-3-methyladenine glycosylase gene, and Allene oxide synthase gene were induced to the highest level within 12 hrs. Subsequently, the expression of five genes decreased at 24 and 48 hrs of treatment except for UDP-glucosyl transferase gene which showed stable expression at all time. Apart from others, dolichyl-diphosphooligosaccharide glycosyl transferase gene was induced to the highest level at 24 hrs and was stable until 48 hrs. This suggested that the induction of genes expression by flooding occurred within 12-24 hrs and most genes in this finding involved in stress responses which were easily upregulated in rheophytic plants during submergence. The more differentially expressed genes should be analysed for a further understanding of the rheophytic habit of this plant.

Analysis on genetic relationship of medicinal species of Dipsacus marked by ISSR, SCoT, and SRAP / 中草药

Objective To detect the genetic relationship among nine medicinal species of Dipsacus in China. Methods The polymorphic bands were detected by three labeling methods (ISSR, SCoT, and SRAP). The genetic distances were calculated by TREECONW software and the systematic diagram of genetic relationship was clustered by UPGMA method. Results The percentage of polymorphism by ISSR, SCoT and SRAP markers showed little difference with value at 90.4%, 88.5%, and 88.2%, respectively, which indicated that the genetic polymorphism of the tested materials was very rich. The genetic distance between Dipsacus chinensis and D. japonicus was the largest, which indicated their genetic relationship was the most distant. Nine medicinal species of Dipsacus were all divided into three groups by three markers, that D. chinensi and D. lijiangensis, D. asperoides and D. japonicas were respectively clustered into one group, D. asperoides var. omiensis was alone clustered, the other four species were clustered into one group. The clustering results labeled by ISSR and SRAP were basically the same with a slight difference between D. daliensis and D. asperoides. Conclusion The clustering results by there markers was between well consistent with the classical classification, which confirmed that molecular markers can be used as one of the effective methods to reveal the genetic relationship among medicinal species of Dipsacus.

Genetic diversity analysis of Pholidota cantonensis in Zhejiang based on SRAP molecular marker technique / 中草药

Objective: To study the genetic diversity of medicinal plant Pholidota cantonensis Rolfe. By SRAP molecular marker technique, and to lay foundations for the protection of this species. Methods: Seven pairs of primers were screened from 81 SRAP primer pairs, and used for SRAP-PCR; The genetic diversity of 68 P. cantonensis materials from five natural populations in Zhejiang Province was applied to analyze. Results: Totally 197 reproducible bands were obtained, and 190 bands of which were polymorphic bands. Each primer pairs contributed 27.29 band on average, and the effective number of allele (Ne) was 1.238 1. At the species level, the percentage of polymorphic bands (PPB) was 96.45%, and the Nei’s genetic diversity (H) and the Shannon’s information index (I) were 0.188 2 and 0.312 5, respectively. At the population level, the PPB ranged from 30.46% to 46.19% with an average PPB of 40%, their H and I were 0.089 1-0.155 9 and 0.138 7 - 0.234 9, with average levels from 0.120 8 to 0.186 4, respectively. The coefficient of gene differentiation (Gst) for five populations was 35.81%, and the gene flow (Nm) was 0.896 3. The Unweighted Pair Group Method with Arithmetic (UPGMA) mean clustering results indicated that 68 samples were divided into five groups, and P. cantonensis plants from the same population were clustered into the same clade. Conclusion: P. cantonensis showed a high level of genetic diversity, and its genetic differentiation as well as gene flow were found among the five populations; However, but the genetic differentiation happened mainly within each population.

Genetic diversity and quality analysis of Rehmannia glutinosa in different germplasm / 中国中药杂志

The study aims at evaluating genetic diversity and medicinal quality of cultivated germplasm in Rehmannia glutinosa, and providing theoretical guidance for screening excellent germplasm. The genetic diversity of 21 species of R. glutinosa were analyzed by SRAP molecular markers, and the catalpol and verbascoside was determined by HPLC. The mass fraction of catalpol and verbascoside in R. glutinosa germplasm were respectively in the range of 2.393%-6.519% and 0.063%-0.478%, the germplasm 14, 16, 15 and 20 germplasm, witch catalpol and verbascoside content was higher. A total of 57 bands were produced by 10 primer, among which 40 polymorphic bands were polymorphic bands, and the percentage of polymorphic loci was 8.77%-54.39%, the Nei's genetic diversity index (H) was 0.374 1, Shannon's polymorphism information index (I) was 0.546 6. Gst and gene flow Nm were 0.608 8 and 0.321 3, respectively. Based on the genetic uniformity, 21 species of germplasm were grouped into 2 categories. The genetic diversity level of R. glutinosa was medium low. The comprehensive consideration of the genetic diversity and the content inculde catalpol and verbascoside, germplasm 7 and germplasm 18 could be used as the preferred materials for the cultivation of reticulum. Germplasm 15 and 16 can be used as the preservation and breeding object of rhubarb germplasm.

Genetic diversity of Trichosanthis Fructus for seeds cultivating / 中草药

Objective To reveal the genetic diversity of cultivating resources of Trichosanthes sp. from different areas. Methods The genetic background of 11 populations (including 30 samples) of Trichosanthis Fructus were researched by both ITS and SRAP molecular markers. The software mega 5.0 and NTSY-pc 2.1 were taken to analysis the ITS sequences and SRAP polymorphic bands data respectively. Results There are obvious mutation in ITS sequences among T. kirilowii and T. rosthornii, ITS sequences could authenticate the different resources in some extent. 165 SRAP bands were amplified by using the optimized 15 primer pairs, and 136 were polymorphic bands, the polymorphic rate was up to 82.4%. The samples were separated into two clusters at the genetic similarity coefficient of 0.18, one cluster gathered the samples of Yuelou Num.8 (T. kirilowii) and the other cluster further separated into three sub-clusters at the coefficient of 0.90, which were mainly T. rosthornii. Conclusion There are several species of Trichosanthes have been used to harvest the seeds in China, and the major one is T. rosthornii. The cultivating resources of T. rosthornii have little difference in genetic backgrounds.

Genetic diversity of Chuanmingshen violaceum by SRAP markers / 中草药

Objective: In order to correctly identify the different germplasm resources of Chuanmingshen violaceum and gain the excellent germplasm resources, SRAP molecular marker was used to analyze the genetic diversity of C. violaceum. Methods: C. violaceum was collected from seven different areas, which included 24 samples, the DNA fingerprint of C. violaceum was constructed with SRAP molecular marker, and the genetic diversity was analyzed. Results: Totally 374 bands were amplified by 37 primer pairs, of which 283 bands were polymorphic, and the polymorphic percentage was 75.67%. The SRAP-based genetic similarity coefficient of all samples ranged from 0.7267 to 0.9239, with a mean of 0.8150. The analysis of molecular variance showed higher percentages of genetic variation within population. All the accessions could be distinguished by SRAP markers. The cluster analysis results showed that 63 accessions were classified into seven groups, which were correlated with the geographical distribution of the accessions to some degree. The accessions from Langzhong and Jintang had higher diversity. Conclusion: There actually exists plentiful genetic diversity among the genetic resources of C. violaceum. SRAP marker is a useful method for analyzing the genetic diversity among C. violaceum accessions.

Optimization of SRAP-PCR reaction systems for Curcuma wenyujin / 中草药

Objective: To research on the most appropriate SRAP-PCR amplication system of Curcuma wenyujin. Methods: The concentration of Mg2+, dNTP, template DNA, primer, and Taq polymerase was optimized by amplifying technology and method of PCR based on isolating genomic DNA from C. wenyujin. Results: The best amplification system for C. wenyujin was as follows: total reaction volume of 25 μL, including Mg2+ (25 mmol/L) 2.0 μL, Taq polymerase(5 U/μL)0.4 μL, random primer (20 μmol/L) 2 μL, template DNA (5 ng/μL) 2.0 μL, dNTP (25 mmol/L) 2.0 μL, 10 × PCR buffer 2.5 μL. Conclusion: The stripes of amplification products are clearness, high brightness, and good reproducibility. It indicates that the reaction system and reaction procedures which are confirmed in this experiment are applied to the SRAP molecular markers in C. wenyujin, based for the genetic diversity of C. wenyujin.

Research on SRAP molecular markers in germplasm resources of Coix lacryma-jobi / 中草药

Objective: To study the 25 germplasm resources of Coix lacryma-jobi by the technique of SRAP molecular markers. Methods: Taking genome DNA in the leaves of C. lacryma-jobi extracted by modified CTAB method as template. Six ideal primer pairs were selected from 88 primer pairs by using the optimized SRAP-PCR reaction system and the amplification of 25 germplasm resources by SRAP-PCR were carried out. Results: The total 66 clear and repeatable polymorphic bands were obtained, and the polymorphic rate was up to 93%. The coefficient of genetic similarity among each species was calculated by using NTSYPC 2.1 software. The results indicated that the coefficient of genetic similarity of these materials ranged from 0.48 to 0.82. Conclusion: The 25 populations of C. lacryma-jobi could be clustered into four groups.

High genetic diversity and small genetic variation among populations of Magnolia wufengensis (Magnoliaceae), revealed by ISSR and SRAP marker

Background Genetic diversity and genetic variation of 10 populations and subpopulations of Magnolia wufengensis, a new and endangered endemic species, were examined by inter simple sequence repeat (ISSR) and sequence-related amplified polymorphism (SRAP) molecular markers. Compared with other endangered endemic Magnolia taxa, M. wufengensis holds a relatively high level of genetic variation. Result Total genetic diversity was found to be 87.7% for ISSR and 88.0% for SRAP markers. For polymorphic loci (P), the effective mean number of alleles (Ae) was 1.414 for ISSR markers and 1.458 for SRAP markers, while the mean expected heterozygosity (H) was 0.256 using ISSR and 0.291 for SRAP markers. Within-population variation was estimated for P as 74.9% using ISSR and 74.6% with SRAP markers; the number of alleles Ae was 1.379 with ISSR and 1.397 for SRAP and H 0.235 with ISSR and 0.247 for SRAP markers. Conclusion The analysis of molecular variation of both ISSR and SRAP marker systems indicated that most genetic variation is within populations, with values of 90.64% and 82.92% respectively. Mantel tests indicated a moderate association between the two marker systems and a low correlation between genetic and geographic distances. High levels of genetic diversity and low levels of population divergence suggest that genetic drift is not currently of great concern for this species. Severe habitat loss and fragmentation, predominantly ascribed to anthropogenic pressures, caused in-situ developing restriction of this species. Action for conserving this rare species for its long-term survival should be taken immediately.

Genetic diversity of Gastrodia elata based on SRAP analysis / 中草药

Objective: In order to correctly identify the different germplasm resources of Gastrodia elata and gain the excellent germplasm resources, SRAP molecular marker was used to analyze the genetic diversity of G. elata. Methods: G. elata was collected from seven different areas, which included 24 smaples of G. elata f. elata, G. elata f. g1auca and G. elata f. viridis, the DNA figerprint of G. elata was constructed with SRAP molecular marker, and the genetic diversity was analyzed. Results: Six huandred Thirty-seven belts were amplified by 33 primers pairs, and the polymorphic percentage was 73.16%. The range of genetic similarity coefficient was 0.404 0-0.908 0, the genetic similarity coefficient among G. elata f. elata was in the range of 0.906 6-0.996 4, and those of G. elata f. g1auca, G. elata f. viridis, and hybrid was in the range of 0.410 4-0.999 6, 0.541 0-0.950 4 and 0.578 2, respectively. They showed small genetic differences. The analysis of molecular variance showed higher percentages of genetic variation within population than those among populations in all species. Populations showed higher genetic structure (FST = 0.33, P < 0.05). In addition, artificial cultivation had influence to the genetic differentiation, but not significantly. So in terms of different cultivation conditions so far had no significant impact on the genetic differentiation of G. elata. Mantel's test has been used to detect the correlation between genetic distance and geographic distance of all sorts of germplasm resources, the result had no significant correlation, and due to the limited number of samples, the result is not representative. Conclusion: SRAP molecular marker method can more effectively reflect the genetic polymorphisms of G. elata. Compared with other two phenotypes, G. elata f. elata is more conservative and has lower genetic diversity. The other two variants have higher genetic diversity.

In vitro mutation induced by ethylmethane sulfonate in Stevia rebaudiana and SRAP identification of salt-tolerant mutants / 中草药

Objective: The in vitro mutation system of Stevia rebaudiana induced by ethylmethane sulfonate (EMS) was established, and the salt-tolerant mutants were identified by SRAP. Methods: S. rebaudiana plantlets were inoculated on MS media containing NaCl with different concentration to screen the salt-tolerant critical concentration. Plantlets were treated with EMS at different concentration and for different time periods, and EMS mutagenized stems were inoculated on MS medium containing critical NaCl concentration to screen the tolerant variants by SRAP markers. Results: The critical salt concentration of S. rebaudiana plantlets was 1.0%, and the suitable concentration and time of EMS were 0.8%-1.0% and 8-10 h. Among the screened 41 S. rebaudiana tolerant mutants by SRAP molecular markers, four were mutated at DNA level, and the mutation rate was 9.76%. Conclusion: The in vitro mutagenesis system of S. rebaudiana with EMS has been established, which provides a new breeding way for high-yield salt-tolerant S. rebaudiana.

Investigation of genetic diversity of yam cultivars by SRAP markers / 中国药学杂志

OBJECTIVE: To investigate the genetic diversity of yam resources in China and provide reliable molecular evidences for cultivar identification and genetic relationship. METHODS: Sequence-related amplified polymorphism (SRAP) was applied to detect the genetic diversity of 21 yam cultivars from eight cultivated populations, Popgene software (version 1.32) was used to calculate genetic parameters, UPGMA clustering was carried out using Mega software (version 4.1), and principal component analysis was performed with the help of Ntsys-pc (Version 2.1) program. RESULTS: Three hundreds and nine bands were amplified by 20 SRAP primer combinations, of which 289 bands were polymorphic. The Nei's gene diversity index (H=0.2881), Shannon's information index (I=0.4416) and total genetic diversity (Ht=0.2891) revealed a relatively high genetic diversity level among yam populations. Within populations, HNWX population showed the highest genetic diversity (PPB= 35.92%), whereas the lowest diversity (PPB=0) was observed in FJSM and ZJRG populations. The genetic differentiation coefficient (Gst=0.8658) and genetic flow (Nm=0.0075) indicated that the genetic diversity among populations was higher than that of within populations. A well-separated groups coupling with cultivated species was formed according to the genetic distance (GD) ranging from 0.0498-0.4879 of the investigated populations. Twenty-one investigated cultivars could be distinguished and be effectively grouped into four origins, i.e., Dioscorea opposita Thunb., Dioscorea alata Linn., Dioscorea persimilis Prain et Burkill. and Dioscorea fordii Prain et Burkill.. CONCLUSION: The genetic diversity level of yam resources is high, and relatively high genetic variation exists among four yam species. SRAP marker is an effective method to identify and classify numerous yam cultivars.

Genetic molecular analysis of Coffea arabica (Rubiaceae) hybrids using SRAP markers

In Coffea arabica (arabica coffee), the phenotypic as well as genetic variability has been found low because of the narrow genetic basis and self fertile nature of the species. Because of high similarity in phenotypic appearance among the majority of arabica collections, selection of parental lines for inter-varietals hybridization and identification of resultant hybrids at an early stage of plant growth is difficult. DNA markers are known to be reliable in identifying closely related cultivars and hybrids. Sequence Related Amplified Polymorphism (SRAP) is a new molecular marker technology developed based on PCR. In this paper, sixty arabica-hybrid progenies belonging to six crosses were analyzed using 31 highly polymorphic SRAP markers. The analysis revealed seven types of SRAP marker profiles which are useful in discriminating the parents and hybrids. The number of bands amplified per primer pair ranges from 6.13 to 8.58 with average number of seven bands. Among six hybrid combinations, percentage of bands shared between hybrids and their parents ranged from 66.29% to 85.71% with polymorphic bands varied from 27.64% to 60.0%. Percentage of hybrid specific fragments obtained in various hybrid combinations ranged from 0.71% to 10.86% and ascribed to the consequence of meiotic recombination. Based on the similarity index calculation, it was observed that F1 hybrids share maximum number of bands with the female parent compared to male parent. The results obtained in the present study revealed the effectiveness of SRAP technique in cultivar identification and hybrid analysis in this coffee species. Rev. Biol. Trop. 59 (2): 607-617. Epub 2011 June 01.

En Coffea arabica (café arabica), el fenotipo y la variabilidad genética son bajos debido a la estrecha base genética y la autofecundación de la especie. Por su alta similitud fenotípica entre la mayoría de las colecciones de arábica, la selección de líneas parentales para hibridación entre variedades, y la identificación de los híbridos resultantes en una fase inicial de crecimiento, es difícil. Para la identificación de variedades estrechamente relacionadas y sus híbridos, los marcadores de ADN son confiables, pero los polimorfismos de amplificación de secuencias relacionadas (SRAP, por sus siglas en inglés) constituyen una nueva tecnología de marcadores moleculares basada en PCR. En este trabajo, sesenta progenies arábica híbridas, pertenecientes a seis cruces, fueron analizadas utilizando 31 marcadores altamente polimórficos. El análisis reveló siete tipos de perfiles de marcadores que son útiles en la discriminación de los progenitores y los híbridos. El número de bandas amplificadas por pares de cebadores estuvo entre 6.13 a 8.58 con un promedio de siete bandas. Entre las seis combinaciones de híbridos, el porcentaje de bandas compartidas entre híbridos y sus progenitores estuvo entre 66.29% y 85.71% con bandas polimórficas que variaron entre 27.64% y 60.0%. El porcentaje de fragmentos híbridos específicos obtenidos en diversas combinaciones híbridas varió entre 0.71% y 10.86% lo que se atribuye a la recombinación meiótica. Con base en el cálculo del índice de similitud, se observó que los híbridos F1 compartieron un número máximo de bandas con el progenitor femenino que con el masculino. Los resultados obtenidos en este estudio muestran la eficacia de la técnica de SRAP en la identificación de cultivos e híbridos de esta especie de café.

Genetic diversity of Lonicera macranthoides geographical populations revealed by SRAP markers / 中草药

Objective: To research genetic diversity of different Lonicera macranthoides geographical populations. Methods: Seventeen materials were estimated by the approach of sequence-related amplified polymorphism (SRAP). The data of amplified bands were analyzed by Popgene 1.31 and Treeconw software. The system diagram of genetic relationship was built by UPGMA. Results: The 24 pairs of SRAP primers combination employed produced a total of 239 discernable and reproducible amplified fragments. Among them there were 210 polymorphic bands. The percentage of polymorphic bands within different populations was 87.8%; Genetic diversity analysis showed that Nei's gene diversity (He) was 0.239 9 and Shannon's genetic diversity index (I) was 0.372 4. Basis on the DNA molecular level, the results indicated that there was abundant genetic diversity among the tested materials. Genetic similarity coefficient ranges changed from 0.442 3 to 0.940 2. The dendrogram including all samples was obtained by UPGMA. In the dendrogram, there were two cluster groups. Conclusion: The genetic diversity of L. macranthoides geopraphical population in China is plentiful. SRAP markers can be effectively applied to genetic analysis in L. macranthoides populations.

Genetic diversity of Liriope muscari using SRAP markers / 中草药

Objective: To study the genetic diversity of Liriope muscari. Methods: The genetic diversity of 47 populations was analyzed by SRAP marker technique. Results: Fifteen primers amplified 323 polymorphic bands, and the average percentage of polymorphic bands reached to 88.47%. The average of polymorphism information content (PIC) was 0.90. Nei's gene diversity index (H) was 0.197 7 and Shannon's information index (I) was 0.319 0. Gene differentiation index (Gst) was 0.252 8. Cluster analysis using UPGMA method showed that genetic similarity coefficient ranged in 0.59-0.99. Conclusion: L. muscari shows higher genetic diversity and the majority of genetic variation occurs in populations.