Amplified Fragment Length Polymorphism Technique Research Articles

Epigenetic architecture of Pseudotaxus chienii: Revealing the synergistic effects of climate and soil variables.

Whether conifers can withstand environmental changes especially temperature fluctuations has been controversial. Epigenetic analysis may provide new perspectives for solving the issue. Pseudotaxus chienii is an endangered gymnosperm species endemic to China. In this study, we have examined the genetic and epigenetic variations in its natural populations aiming to disentangle the synergistic effects of climate and soil on its population (epi)genetic differentiation by using amplified fragment length polymorphism (AFLP) and methylation-sensitive AFLP (MSAP) techniques. We identified 23 AFLP and 26, 7, and 5 MSAP outliers in P. chienii. Twenty-one of the putative adaptive AFLP loci were found associated with climate and/or soil variables including precipitation, temperature, K, Fe, Zn, and Cu, whereas 21, 7, and 4 MSAP outliers were significantly related to precipitation of wettest month (Bio13), precipitation driest of month (Bio14), percent tree cover (PTC), and soil Fe, Mn, and Cu compositions. Total precipitation and precipitation in the driest seasons were the most influential factors for genetic and epigenetic variation, respectively. In addition, a high full-methylation level and a strong correlation between genetic and epigenetic variation were detected in P. chienii. Climate is found of greater importance than soil in shaping adaptive (epi)genetic differentiation, and the synergistic effects of climate and climate-soil variables were also observed. The identified climate and soil variables should be considered when applying ex situ conservation.

Molecular Characterization of Olive (Olea europaea L.) Cultivars in Duhok Governorate

The olive tree (Olea europaea L.) was considered one of the most cultivated crops in the Mediterranean region, and it had an economically important value due to it began consumed as a table olive and oil production. In this study, AFLP and SRAP techniques were used to identify 15 cultivars of olive plants that were collected from 12 different locations in the Duhok governorate, eleven primer combinations of sequence-related amplified polymorphism (SRAP) and ten primer combinations of amplified fragment length polymorphism (AFLP) were used to fingerprinting these cultivars. The results show that the SRAP marker had the least genetic distance 0.241between (Shtla falay olive and Khodeiri olive) and the highest value 0.946 was found between (Zawita olive and Sorani olive). While AFLP marker show the lowest value of 0.107 between (Nzarky olive and Dger olive) and the highest value of 0.916 between (Amedi olive and Shtla falay olive). Cluster analysis based on AFLP and SRAP combined data clearly differentiated the genotype and it showed it was a powerful tool to evaluate the genetic distance and fingerprinting of these tested cultivars. Establishing the genetic relationships between cultivars and species, and the construction of genetic maps, had the main aims for the researchers in plant breeding programs and to understand the plant evolution lingues, and as a conclusion, this study demonstrates that AFLP and SRAP markers both are powerful tools in plants tree genetics.

In vitro propagation for conservation and genetic fidelity of the near threatened Dimocarpus longan plant

BackgroundDimocarpus longan is a tropical tree that produces edible fruit. It is a neglected plant species that is listed as near threatened. In spite of its economic value, the propagation of longan cultivar using conventional methods is extremely difficult. The goal of this research is to produce and conserve this plant through in vitro propagation. ResultsIn order to form new shoots, sterilized shoot tip explants were cultured on Murashige and Skoog (MS) medium supplemented with benzyl adenine (BA) or 2-isobentenyl-adenine (2ip). For direct organogenesis, young leaves of new shoots were cultured on MS medium fortified with various concentrations of Thidiazuron (TDZ) or 6-(4-Hydroxy-3-methylbut-2-enylamino purine) (Zeatin). Gibbrellic acid (GA3) at different levels alone or in combination was used for shoot elongation. Also, indole-3-butyric acid (IBA) and naphthalene acetic acid (NAA) were used for root formation. MS medium supplemented with 1.00 mg/l 2ip was suitable for inducing axillary shoots from shoot tips (4.0 axillary shoots/explant). The highest significant 76% and numbers of adventitious buds from leaf base were achieved on MS medium containing 1.0 mg/l TDZ. These buds developed into the longest plantlets on GA3 at 3.0 mg/l and rooted well in ½MS containing 1.50 mg/l IBA plus 0.50 mg/l (NAA). About 70% in vitro plants were successfully acclimatized. The AFLP profile illustrated the genetic stability of gene expression action. The amplified fragment length polymorphisms (AFLPs) profile illustrated the progenies were extremely similar to the mother plants. According to our findings, MS medium containing 25 ppm salicylic acid (SA) and 5 ppm methyl jasmonate (MeJA) produced the highest percentage of apigenin in longan calli (77.09 and 2.637%, w/w). ConclusionA successful and efficient micropropagation protocol has been developed and described here for the first time, and it will be very useful for the clonal propagation and conservation of the near-threatened Dimocarpus longan plant. Micropropagated plants are genetically identical to the donor plant using the AFLP technique. The usefulness of salicylic acid and methyl jasmonate as elicitors for increasing in vitro production of secondary metabolites in plants is demonstrated in this work.

AFLP analysis of Genetic diversity and Phylogenetic relationships of Vigna radiata (L) Wilczek

Among the important pulse-yielding plants Vigna radiata is important. In the Indian subcontinent, it is an essential source of dietary protein particularly in the vegetarian population. It contains abundant nutrients with biological activities. Soaked seeds are eaten as a good source of protein and mature seeds are cooked or can be sprouted for human consumption. Mung beans and its sprouts contain chemical constituents like flavonoids, phenolic acids, organic acids, amino acids, carbohydrates, and lipids. It is related to different biological activities like antioxidant, antimicrobial, antitumor, anti-inflammatory, anti-diabetic, and antihypertensive effects, etc. The seeds are a traditional source of cures for paralysis, rheumatism, coughs, fevers, and liver ailments. As a green manure or cover crop, the plant can be grown, enriching the soil with the nitrogen formed on its roots. The powerful Amplified Fragment Length Polymorphism (AFLP) is a molecular marker used for a variety of applications like assessing genetic diversity within species or among closely related species, inferring population-level phylogenies, generating genetic maps, and determining the relatedness among cultivars. AFLP has become extremely beneficial in the study of taxa including bacteria, fungi, and plants, where much is still unknown about the genomic makeup of various organisms. A study was made to assess the genetic diversity and phylogenetic relationships of a set of five varieties of Vigna radiata (L) Wilczek using the AFLP technique. Five different varieties of Vigna radiata viz. B1, TARM 2, PDM 84, TM 99, and TM 98 were subjected to AFLP analysis. A total of 471 fragments were scored across all the 12 AFLP primer sets used and the results were used to plot a dendrogram. It was observed that these five varieties formed three distinct groups among which the aromatic B1 variety formed a distinct group.

Clinical profile, antifungal susceptibility, and molecular characterization of Candida auris isolated from patients in a South Indian surgical ICU.

BackgroundCandida auris is an emerging multidrug resistant yeast which causes blood stream infection especially among critically ill patients. This yeast can also colonize patients and are isolated from hospital environment causing outbreaks in hospital settings. ObjectiveTo describe possible outbreak of C. auris infection in surgical ICU and characterize the isolates by molecular typing and azole resistance mechanism. MethodsAfter isolation of Candida auris from cluster of patients from surgical ICU, environment survey was done to identify the source in the hospital. The identity of the isolates was confirmed by Matrix Assisted Laser Desorption Ionisation Time of Flight mass spectroscopy and sequencing 26S and ITS region of rDNA. Molecular typing was done by fluorescent amplified fragment length polymorphism technique. Antifungal susceptibility testing was performed by CLSI broth dilution technique. ERG11 gene was sequenced to screen for mutations responsible for azole resistance. Results and conclusionA total of eight C. auris was isolated during the four months (December 2018–March 2019) suggesting possible of outbreak in surgical ICU of tertiary care center in South India. C. auris (n = 8) was isolated from urine (n = 4), blood (n = 3) and ear discharge (n = 1) samples. Based on 26S sequence analysis all our isolates belonged to South Asian clade. All the isolates had minimum inhibitory concentration (MIC) of ≥16 µg/ml to fluconazole. ERG11 sequence exhibited amino acid substitution Y132F in all the isolates. The two environmental isolates clustered closely with an isolate from urine sample. Adherence to strict infection control practices prevented further spread of infection.

Evaluation of genetic stability in olive callus-induced and meristem-induced shoots using flow cytometry and amplified fragment length polymorphism techniques

BackgroundIn vitro culture of olive, as an economically valuable tree, has fundamentally a genotype-dependant low micropropagation rate which needs to be improved in already established and newly released cultivars. Various plant tissue culture media, planting systems and growth factors were evaluated in two promissing Iranian olive cultivars ˈAminˈ and ˈMeshkatˈ and the commercial Spanish cultivar ˈArbequinaˈ.ResultsThe results showed that cultivars have their specific optimal media, i.e. ˈAminˈ in the MS with 4 mg/L zeatin, ˈArbequinaˈ in the OM with 1 mg/L zeatin, and ˈMeshkatˈ in the OM and MS with 2 mg/L zeatin, which produced significantly a higher number of axillary shoots than other media. The results also indicated a significant improvement in the growth indices of ˈAminˈ (number of axillary shoots) when cultured using periodical mini bioreactor (PMB) in the VS medium. In comparison with VS, OM did not reveal any significant differences when both culturing systems (PMB and semi-solid media (SSM)) were used.Regarding the effect of carbon source and light intensity, mannitol and 2000 cd sr m−2 greatly enhanced ˈArbequinaˈ growth indices (main shoot length and growth quality). The results of genetic stability of callus induced shoots (CIS) and meristem induced shoots (MIS) revealed that 2C DNA value assessed by partec flow cytometery (FCM) had 0.01, 0.03 and 0.08 pg discrepencies in ˈAminˈ, ˈArbequinaˈ and ˈMeshkatˈ, repectively. The Amplified Fragment Length Polymorphism (AFLP) results also indicated that the cultivars were classified regardless of the micropropagation origin (CIS or MIS), except for ˈArbequinaˈ. The AFLP findings showed that ˈArbequinaˈ had the highest dispersal (7–38%) in CIS and MIS, while the Iranian cultivar of ˈMeshkatˈ (5–9%) had the highest stability.ConclusionsThis study indicated the importance of in vitro growth parameters for improving the micropropagation indices of olive cultivars. It showed that optimized protocols (OM, PMB, zeatin, mannitol and 2000 cd sr m−2) co-produced larger calli resulting in indirect organogenesis. Based on FCM and AFLP analysis, it can be concluded that true-to-typeness of micropropagated olive was cultivar-dependent.

Voriconazole resistance genes in Aspergillus flavus clinical isolates

ObjectiveThe present study was designed to discover novel biomarkers involved in voriconazole resistance in clinical isolates of Aspergillus flavus. Materials and methodsTwo voriconazole non-wild-type and two voriconazole-wild-type A. flavus clinical isolates were selected to evaluate possible molecular mechanism involved in A. flavus resistance to voriconazole using the mutation assessment, Quantitative real- time PCR of cyp51A and cyp51C genes and complementary DNA- amplified fragment length polymorphism technique. ResultsNo mutations were seen in the cyp51A and cyp51C genes in voriconazole non-wild-type isolates compared to wild- type and reference strains. Regarding to mRNA expression results, no changes were observed in expression fold of cyp51A and cyp51C mRNA expression level in first non- wild- type isolate compared to wild-type isolate. For second isolate cyp51C mRNA expression level was down regulated (5.6 fold). The set of genes including ABC fatty acid transporter XM- 002375835 and aldehydereductase XM- 002376518 and three unknown functional genes were identified. Based on results, the over-expression of AKR1 and ABC fatty acid transporter in the voriconazole non- wild- type isolates suggests these genes could represent a novel molecular marker linked to the voriconazole resistance in A. flavus. ConclusionThe results obtained in this study showed a novel finding as the authors identified AKR1 and ABC fatty acid transporter genes as possible voriconazole target genes in Iranian clinical isolates of A. flavus.

Genetic diversity analysis of some Egyptian Origanum and Thymus species using AFLP markers

BackgroundIn the present investigation, two genera of family Labiatae (genus Origanum and genus Thymus including their available species in Egypt) were analyzed genetically on DNA level using amplified fragment length polymorphism (AFLP) markers. Four selective primer combinations (E-AGC/M-CAA, E-AGC/M-CAC, E-AGG/M-CTC, and E-ACC/M-CAT) were used to detect genetic variations (polymorphisms) within and between the studied plant species of each genus and with their cultivated relatives. ResultsA total of 193 amplified fragments were obtained overall primer combinations with a disparity in the number of bands for each primer combination. The primer combination E-AGG/M-CTC exhibited a large number of amplicons (67) compared to the other primers with polymorphism percentage 69%. The polymorphism information content (PIC) was calculated for the four primer combinations showing a very high values ranging between 0.98 and 0.99. On the other hand, Thymus species (Thymus vulgaris, Thymus capitatus L., and Thymus decassatus Benth.) exhibited a total number of 171 amplicons for all primer combinations with an average of 42 bands. The primer combination E-AGG/M-CTC produced the largest number of bands (62 bands) with polymorphism percentage 35%, even though the primer combination AGC/M-CAA was more efficient to give high polymorphisms within Thymus species where it resulted in a total of 35 bands with polymorphism percentage 63%. The PIC values were calculated ranging from 0.96 to 0.99. ConclusionOne can conclude that AFLP technique was informative and efficient technique to give a good coverage of Origanum and Thymus genomes. Furthermore, it was helpful to elucidate the genetic variations and phylogenetic relationships within the studied species as a basis for further studies on these genera and related species.

Numerical analysis of phenotypic properties, genomic fingerprinting, and multilocus sequence analysis of Bradyrhizobium strains isolated from root nodules of Lembotropis nigricans of the tribe Genisteae

PurposeThe aim of this study was to estimate the level of genomic and phenotypic diversity as well as the genus and species position of bacterial strains isolated from root nodules of Lembotropis nigricans (family Fabaceae).MethodsThe genomic diversity of studied L. nigricans nodule symbionts was examined by using BOX-PCR and AFLP (amplified fragment length polymorphism) fingerprinting techniques. To assign bacteria to the genus, numerical analysis of phenotypic features and comparative analysis of 16S rDNA sequences were performed. The comparative analysis of combined atpD, dnaK, gyrB, and rpoB gene sequences (multilocus sequence analysis, MLSA) was used to determine the most closely related species to the studied bacteria.ResultsBoth BOX-PCR and AFLP techniques revealed a high level of genomic heterogeneity of L. nigricans nodulators. Among 33 studied bacteria, 32 genotypes were delineated by the AFLP method and 27 genotypes were identified by the BOX-PCR fingerprinting. The numerical analysis of 86 phenotypic characteristics of L. nigricans nodule isolates and reference rhizobia showed that studied bacteria belong to the genus Bradyrhizobium. Affiliation of L. nigricans nodule isolates to the genus Bradyrhizobium was supported by comparative analysis of 16S rDNA sequences and the concatenation of atpD, dnaK, gyrB, and rpoB gene sequences. MLSA indicated also that L. nigricans microsymbionts are members of Bradyrhizobium japonicum.ConclusionL. nigricans root nodule symbionts are members of Bradyrhizobium japonicum and exhibit high phenotypic and genomic diversity important for their survival in soil.

Sequence capture using AFLP-generated baits: A cost-effective method for high-throughput phylogenetic and phylogeographic analysis.

Target sequence capture is an efficient technique to enrich specific genomic regions for high‐throughput sequencing in ecological and evolutionary studies. In recent years, many sequence capture approaches have been proposed, but most of them rely on commercial synthetic baits which make the experiment expensive. Here, we present a novel sequence capture approach called AFLP‐based genome sequence capture (AFLP Capture). This method uses the AFLP (amplified fragment length polymorphism) technique to generate homemade capture baits without the need for prior genome information, thus is applicable to any organisms. In this approach, biotinylated AFLP fragments representing a random fraction of the genome are used as baits to capture the homologous fragments from genomic shotgun sequencing libraries. In a trial study, by using AFLP Capture, we successfully obtained 511 orthologous loci (>700,000 bp in total length) from 11 Odorrana species and more than 100,000 single nucleotide polymorphisms (SNPs) in four analyzed individuals of an Odorrana species. This result shows that our method can be used to address questions of various evolutionary depths (from interspecies level to intraspecies level). We also discuss the flexibility in bait preparation and how the sequencing data are analyzed. In summary, AFLP Capture is a rapid and flexible tool and can significantly reduce the experimental cost for phylogenetic studies that require analyzing genome‐scale data (hundreds or thousands of loci).

AFLP-AFLP in silico-NGS approach reveals polymorphisms in repetitive elements in the malignant genome.

The increasing interest in exploring the human genome and identifying genetic risk factors contributing to the susceptibility to and outcome of diseases has supported the rapid development of genome-wide techniques. However, the large amount of obtained data requires extensive bioinformatics analysis. In this work, we established an approach combining amplified fragment length polymorphism (AFLP), AFLP in silico and next generation sequencing (NGS) methods to map the malignant genome of patients with chronic myeloid leukemia. We compared the unique DNA fingerprints of patients generated by the AFLP technique approach with those of healthy donors to identify AFLP markers associated with the disease and/or the response to treatment with imatinib, a tyrosine kinase inhibitor. Among the statistically significant AFLP markers selected for NGS analysis and virtual fingerprinting, we identified the sequences of three fragments in the region of DNA repeat element OldhAT1, LINE L1M7, LTR MER90, and satellite ALR/Alpha among repetitive elements, which may indicate a role of these non-coding repetitive sequences in hematological malignancy. SNPs leading to the presence/absence of these fragments were confirmed by Sanger sequencing. When evaluating the results of AFLP analysis for some fragments, we faced the frequently discussed size homoplasy, resulting in co-migration of non-identical AFLP fragments that may originate from an insertion/deletion, SNP, somatic mutation anywhere in the genome, or combination thereof. The AFLP–AFLP in silico–NGS procedure represents a smart alternative to microarrays and relatively expensive and bioinformatically challenging whole-genome sequencing to detect the association of variable regions of the human genome with diseases.

Analysis of genetic diversity evaluation of Lysiphyllum strychnifolium (Craib) A. Schmitz in Thailand using amplified fragment length polymorphism markers

Lysiphyllum strychnifolium (Craib) A. Schmitz, a Leguminosae-Caesalpiniodeae species, is colloquially known in Thai as Ya Nang Daeng. In Thai traditional medicine, the leaves, stems and roots have been used to relieve fever and alcohol toxication. The stem has also been used for anti-HIV-1 integrase, anti-allergy and anti-cancer treatment. Confusion over herbal drugs has arisen from the homonymity and morphological characteristics, which present many similarities within its family. Thus, germplasm collection and genetic diversity analysis are prerequisites for a breeding program. In this study, the amplified fragment length polymorphism (AFLP) marker was used to identify and elucidate the phylogenetic relationships among 98 accessions of Ya Nang Daeng throughout Thailand. Thirteen AFLP primer combinations generated 379 bands, of which 359 were polymorphic. The value of the similarity coefficient varied from 0.52 to 0.93, with a mean of 0.73. A phylogenetic tree was constructed using the unweighted major clusters at a cut-off genetic mean (UPGMA) clustering method. The phylogenetic tree derived from the AFLP data showed that the Ya Nang Daeng accessions could be divided into two clusters. Cluster 1 could be further classified into five subgroups, while a similarity coefficient of 0.23 could be used to separate an outgroup from Ya Nang Daeng. The result of principal coordinate analysis and morphological grouping corresponded to the UPGMA clustering. This study showed that the AFLP technique is a useful tool for estimating DNA polymorphism of the genetic diversity and differentiation of related plants. This genetic information will provide useful information for a breeding program in the future.

Before-after analysis of genetic and epigenetic markers in garlic: A 13-year experiment

Fil: Gimenez, Magali Diana. Consejo Nacional de Investigaciones Cientificas y Tecnicas. Centro Cientifico Tecnologico Conicet - San Juan. Centro de Investigaciones de la Geosfera y Biosfera. Universidad Nacional de San Juan. Facultad de Ciencias Exactas Fisicas y Naturales. Centro de Investigaciones de la Geosfera y Biosfera; Argentina

OPTIMATION OF DNA EXTRACTION PROTOCOL OF Elaeidobius kamerunicus

African pollination weevil (Elaeidobius kamerunicus Faust) has an important role in the productivity of Indonesian oil palm plantation. Up to now, there has not been a comprehensive biological study of the species at molecular level. The basic knowledge is very useful for exploitation of the weevil for effective oil palm fruit set development. This research aimed to obtain DNA extraction protocol of E. kamerunicus for DNA fingerprinting of the species. Results showed that using a DNA extraction kit,material disruption by using micro pestle resulted the highest quantity of DNA, while there were no significant differences of resulted DNA quantity among treatments using tissue lyser for material disruption. DNA extracted by using micro pestle or tissue lyser for material disruption is adequate for DNA fingerprinting using AFLP (Amplified Fragment Length Polymorphism) and sequencing techniques.

Screening for single nucleotide polymorphisms in highly degraded DNA by using the amplified fragment length polymorphism technique

Short tandem repeat (STR) analysis is generally used for human identification of forensic samples; however, standard STR analysis sometimes fails to generate full profiles since DNA is frequently degraded by various environmental factors. Recently, single nucleotide polymorphism (SNP) analysis has attracted attention for human identification since the shorter amplicons are better suited for degraded samples. Though various SNP loci are used for analysis of degraded samples, it is unclear which ones are more appropriate. To identify SNPs that were resistant to degradation, we artificially degraded DNA obtained from the buccal swabs of six volunteers and the K562 cell line by heat treatment. Subsequently, the amplified fragment length polymorphism (AFLP) technique was used for SNP screening. We focused on the AFLP bands detected in both the heat-treated and untreated samples, and DNA extracted from these bands was directly sequenced. DNA degradation increased as the duration of heat treatment increased, and no STR profiles could be generated after 6h of heat treatment. When the AFLP band patterns were compared between 6h heat-treated and untreated samples, eight common bands were detected. The sequences of the DNA fragments of these common bands had higher adenine-thymine (A-T) content and included 17 SNPs. The SNPs detected in the heat-treated and untreated samples were considered to be resistant to degradation. Although there was a little information available in databases regarding the nine SNPs identified in this study, this study shows that some of these SNPs might be useful for human identification of extremely degraded DNA.

OPTIMATION OF DNA EXTRACTION PROTOCOL OF Elaeidobius kamerunicus

African pollination weevil (Elaeidobius kamerunicus Faust) has an important role in the productivity of Indonesian oil palm plantation. Up to now, there has not been a comprehensive biological study of the species at molecular level. The basic knowledge is very useful for exploitation of the weevil for effective oil palm fruit set development. This research aimed to obtain DNA extraction protocol of E. kamerunicus for DNA fingerprinting of the species. Results showed that using a DNA extraction kit,material disruption by using micro pestle resulted the highest quantity of DNA, while there were no significant differences of resulted DNA quantity among treatments using tissue lyser for material disruption. DNA extracted by using micro pestle or tissue lyser for material disruption is adequate for DNA fingerprinting using AFLP (Amplified Fragment Length Polymorphism) and sequencing techniques.

Assessment of Genetic Relationships in Breeding Lines and Cultivars of Brassica napus and Their Implications for Breeding Winter Oilseed Rape

The level of genetic diversity or similarity between parents has been proposed as a predictor of F1 generation performance and heterosis for seed yield. The objective of this study was to determine the feasibility of using amplified fragment length polymorphism (AFLP) fluorescent‐labeled primers to evaluate the genetic relationships between 101 oilseed rape (Brassica napus L.) genotypes to maximize the selection of different parents in a breeding program. The collection of genotypes included 25 cytoplasmic male sterility ogura and 21 restorer lines (Rfo). The set of genotypes was completed with B. napus cultivars of Polish, European, and Asian origin and resynthesized lines. Ten primer‐enzyme combinations (EcoRI and MseI) were used. Genetic similarities among genotypes were estimated in accordance with the Nei and Li coefficient of similarity, and then the unweighted pair group method using arithmetic averages method was applied for the creation of a dendrogram. A total of 344 reproducible, polymorphic markers were obtained. Unweighted pair group method using arithmetic averages clustering based on the Nei and Li coefficient of similarity separated the 101 genotypes into four distinct clusters. Analysis of molecular variance showed that 90% of the detected variation was found within the group, while the variation among groups contributed to only 10%. Our results indicate that the AFLP technique seemed to be a suitable molecular method for assessing genetic relationships between oilseed rape inbred lines with high levels of accuracy. Genetic similarity evaluated on the basis of AFLP molecular markers is a significant addition to classical methods of selection of parental components of winter oilseed rape hybrid varieties (e.g., ‘Poznaniak’).

Polymorphic CAG Repeat Numbers in the Androgen Receptor Gene of Female Pattern Hair Loss in a Han Chinese Population

Objective: To investigate the association of CAG repeat numbers in the androgen receptor (AR) gene with female pattern hair loss (FPHL) in a Chinese population. Methods: A total of 200 Han Chinese patients with FPHL (142 Ludwig II and 58 Ludwig III cases) and 200 healthy controls were enrolled in this study. The polymorphism of CAG repeat numbers was analyzed by the fluorescent amplified fragment length polymorphism technique. Results: The CAG biallelic mean length was 23.73 ± 2.04 repeats in Han Chinese FPHL patients and 23.90 ± 2.13 repeats in healthy controls, without any significant difference between the two groups (p = 0.481). In addition, neither the shorter nor the longer CAG repeat numbers were significantly different between FPHL and control subjects (p = 0.726, p = 0.383). Conclusion: The polymorphism of CAG repeat numbers of the AR gene may not be the genetic marker of FPHL in a Chinese population.

Genetic diversity and variability in populations of the white wax insect Ericerus pela, assessed by AFLP analysis.

The white wax insect Ericerus pela Chavannes (Hemiptera: Coccoidea) is an economically valuable insect species that has been used for over a thousand years in China. The present study focuses on assessing the genetic variability in different populations of E. pela collected from seven Chinese provinces. The amplified fragment length polymorphism technique was used to generate DNA fingerprints of individuals from each population using nine primer combinations (EcoRI-MseI). A total of 435 polymorphic loci were generated; fragment sizes ranged from 200 to 1000 bp. The percentage of polymorphic loci was 85.29%. Nei's genetic diversity and Shannon index indicated consistency in the results, which showed that the Sichuan population had the highest diversity, followed by Yunnan and Zhejiang populations. Dendrogram analysis showed the shortest genetic distance between the Sichuan and Yunnan populations, suggesting that they probably form sister groups. High genetic differentiation between population values among all sampled populations indicated a low degree of genetic variability within each population (40.85%) and higher variation among populations (59.15%). Gene flow estimate values were low in all samples, suggesting low gene flow from events such as interbreeding and migration. Low gene flow values also suggested that populations among species of E. pela might become genetically heterogeneous, due to counteracting forces such as strong differential selection. Our data support the probability that E. pela will remain localized, and has a low potential to spread beyond current habitats.

Weak isolation by distance in Diaperis boleti, a fungivorous saproxylic beetle.

Living in unstable habitats is expected to decrease the intensity of isolation by distance in populations through the need for frequent movements of individuals. Insects associated with fruiting bodies of fungi therefore are supposed to have weak spatial genetic structure of populations compared with those living in more stable habitats. With the use of an amplified fragment length polymorphism technique, this study investigated the isolation by distance, inbreeding, and genetic diversity in Diaperis boleti (L.) (Coleoptera: Tenebrionidae), a fungivorous saproxylic beetle that inhabits sporocarps of Laetiporus sulphureus (Bulliard) Murrill (Polyporales) on trees growing in highly-fragmented agricultural landscapes. Isolation by distance was tested with spatial autocorrelation analysis of kinship (individual-based approach) and correlating matrices of genetic and geographic distances with the Mantel test (population-based approach). These results were compared with the results obtained for saproxylic beetles living in the same landscape but differing in ecological preferences. It was shown that the species dependent on sporocarps of wooddecomposing fungi had higher variability, lower individual inbreeding, and less intensive isolation by distance pattern than saproxylic beetles living in tree hollows. It was also demonstrated that spatial autocorrelation analysis of kinship is a more sensitive approach for detecting finescale spatial genetic structure than the Mantel test.