Perfect Repeats Research Articles

Characterization and novel Est-SSR marker development of an important Chinese medicinal plant, Morinda officinalis How (Rubiaceae)

Morinda officinalis How (Rubiaceae) is well known as one of the four most popular medicinal plants in the southern region of China. In order to assess its genetic diversity and assist the development of its molecular assisted-selection breeding in China, we constructed the transcriptome database of M. officinalis using fresh leaf material and developed novel expressed sequence tag–simple sequence repeat (EST-SSR) markers for this important medicinal plant species. Our study identified 8064 potential EST-SSRs from 6668 unigenes. For perfect repeat motifs, the mononucleotide repeats were most abundant (4221; 52.34%), followed by the trinucleotide repeats (1588; 19.69%), and the dinucleotide ones (1040; 12.90%). The seven most commonly occurring repeat motifs were A/T (4661), AG/CT (588), AAG/CTT (408), AC/GT (353), ATC/ATG (338), AT/AT (296) and AGG/CCT (277). Of a total of 5605 SSR primer sets that were successfully designed de novo, 100 were selected for validation analysis. There, 24 primer sets were polymorphic across the 96 M. officinalis individuals collected from four different populations in Guangdong province. Based on cross-species transferability test, 19 and 22 primer sets produced successful amplification in M. umbellata subsp. obovata and M. parvifolia, respectively. These novel EST-SSR markers with high cross-species transferability between M. officinalis and its closely-related species could serve as aids to future studies in genetic diversity, conservation, assisted-selection breeding and genes associated with the medical uses of the genus.

Imperfect and Compound Microsatellites in the Genomes of Burkholderia pseudomallei Strains

Evolution of microsatellites (or simple sequence repeats, SSRs) is a complex process that converts perfect repeats to novel structural elements with functions poorly understood, such as imperfect and compound microsatellites. An in silico analysis often Burkholderia pseudomallei genomes revealed 215683 micro-satellites, and more than 98% of them proved imperfect. The density of microsatellites in the genome ranged from 2922.7 to 3022.6 per Mbp. Approximately 10.20-10.67% of the repeats were parts of compound micro-satellites. The of compound microsatellite density varied from 144.7 to 150.6 per Mbp. Between-strain differences in microsatellite distribution were explained by a direct correlation of the SSR density with the GC content and an inverse relationship between the SSR density and the genome size. For each B. pseudomallei chromosome, the SSR density similarly correlated with its size and GC content. Chromosome 2 showed a significant correlation between the SSR and compound microsatellite densities (r = 0.93, p < 10^(-3)). The association of imperfect and compound microsatellite densities with the structural features of each chromosome and the fact that motifs are degenerate and occur in few copies in the majority of B. pseudomallei microsatellites agree with the previous hypothesis of negative selection affecting extended SSRs. The mechanism of selection possibly involves an accumulation of point mutations, which lead to an interruption of the repeat during replication because easily passable secondary structures may form to stabilize the microsatellite length.

MilR2, a novel TetR family regulator involved in 5-oxomilbemycin A3/A4 biosynthesis in Streptomyces hygroscopicus

Milbemycins produced by several Streptomyces species are a group of 16-membered macrolides with potent insecticidal and anthelminthic activity. Milbemycin A3/A4, the main components of the milbemycins biosynthetic pathway, and 5-oxomilbemycin A3/A4, the analogs of milbemycin A3/A4 without the reduction of the C-5 keto group, have been developed as acaricides, insecticides, and anthelmintics. However, so far, little is known about the regulation of milbemycins biosynthesis, which has greatly hampered the generation of high producing strains by metabolic engineering. Herein, a TetR family regulator MilR2 (encoded by sbi_00792) was identified being involved in activation of 5-oxomilbemycin A3/A4 biosynthesis in a high 5-oxomilbemycins-producing strain Streptomyceshygroscopicus SIPI-KF. TheΔmilR2 mutant with an in-frame deletion of theMilR2 DNA-binding domain resulted in significantly reduced 5-oxomilbemycin A3/A4 production (approximately 36.9 and 39.7%) at tested two time points, and accordingly introduction of an extra copy of milR2 into SIPI-KF led to enhanced production by 12.6 and 34.4%. We further showed that MilR2 could directly repress the transcription of the gene sbi_00791 encoding a putative hydrolase, which is located divergently from milR2. The precise MilR2-binding site consisting of a 7-nt perfect inverted repeat separated by 10-nt (5'-ACCAACCAGCTGGTAAGGGTTGGT-3') was defined. In situ mutagenesis of the MilR2-binding site resulted in 19.7 and 13.5% decreases in 5-oxomilbemycin A3/A4 production, which is much lower than the decreased rates of ΔmilR2. Collectively, the results demonstrated that MilR2 serves as an activator for 5-oxomilbemycin A3/A4 production and the function of MilR2 is only partially mediated through its repression on the transcription of sbi_00791.

The Prevalence and Evolutionary Conservation of Inverted Repeats in Proteobacteria.

Perfect short inverted repeats (IRs) are known to be enriched in a variety of bacterial and eukaryotic genomes. Currently, it is unclear whether perfect IRs are conserved over evolutionary time scales. In this study, we aimed to characterize the prevalence and evolutionary conservation of IRs across 20 proteobacterial strains. We first identified IRs in Escherichia coli K-12 substr MG1655 and showed that they are overabundant. We next aimed to test whether this overabundance is reflected in the conservation of IRs over evolutionary time scales. To this end, for each perfect IR identified in E. coli MG1655, we collected orthologous sequences from related proteobacterial genomes. We next quantified the evolutionary conservation of these IRs, that is, the presence of the exact same IR across orthologous regions. We observed high conservation of perfect IRs: out of the 234 examined orthologous regions, 145 were more conserved than expected, which is statistically significant even after correcting for multiple testing. Our results together with previous experimental findings support a model in which imperfect IRs are corrected to perfect IRs in a preferential manner via a template switching mechanism.

Characterization of 17 novel microsatellite loci for the brown spiny lobster Panulirus echinatus Smith, 1869 using MiSeq

Using high-throughput sequencing technology 17 polymorphic microsatellites with perfect tetra-nucleotide repeats were identified for the brown spiny lobsterPanulirus echinatusSmith, 1869. Two to ten alleles were detected per locus across 30 samples analysed from the Canary Archipelago. Observed and expected heterozygosities per locus ranged between 0.100 and 0.867 and 0.095 and 0.799, respectively. No significant linkage disequilibrium was found between pairs of loci, and only one locus (Pe-L43) deviated from Hardy-Weinberg equilibrium, likely due to the presence of null alleles. Despite its economic importance and the threat of overfishing, population genetics studies of this species are lacking. Therefore, these 17 novel microsatellites markers will be a useful genetic resource for future conservation studies ofP. echinatus.

Physiological characterization, transcriptomic profiling, and microsatellite marker mining of Lycium ruthenicum.

Lycium ruthenicum is a perennial shrub species that has attracted considerable interest in recent years owing to its nutritional value and ability to thrive in a harsh environment. However, only extremely limited transcriptomic and genomic data related to this species can be found in public databases, thereby limiting breeding research and molecular function analysis. In this study, we characterized the physiological and biochemical responses to saline-alkaline mixed stress by measuring photochemical efficiency, chlorophyll content, and protective enzyme activity. We performed global transcriptomic profiling analysis using the Illumina platform. After optimizing the assembly, a total of 68 063 unique transcript sequences with an average length of 877 bp were obtained. Among these sequences, 4096 unigenes were upregulated and 4381 unigenes were down-regulated after saline-alkaline mixed treatment. The most abundant transcripts and over-represented items were assigned to gene ontology (GO) terms or Kyoto Encyclopedia of Genes and the Genomes (KEGG) categories for overall unigenes, and differentially expressed unigenes were analyzed in detail. Based on this set of RNA-sequencing data, a total of 9216 perfect potential simple sequence repeats (SSRs) were identified within 7940 unigenes with a frequency of 1/6.48 kb. A total of 77 primer pairs were synthesized and examined in wet-laboratory experiments, of which 68 loci (88.3%) were successfully amplified with specific products. Eleven pairs of polymorphic primers were verified in 225 individuals from nine populations. The inbreeding coefficient and the polymorphism information content value ranged from 0.011 to 0.179 and from 0.1112 to 0.6750, respectively. The observed and expected heterozygosities ranged from 0.064 to 0.840 and from 0.115 to 0.726, respectively. Nine populations were clustered into three groups based on a genetic diversity study using these novel markers. Our data will be useful for functional genomic investigations of L. ruthenicum and could be used as a basis for further research on the genetic diversity, genetic differentiation, and gene flow of L. ruthenicum and other closely related species.

In silico identification and characterization of stress and virulence associated repeats in Salmonella

So much genomic similarities yet causing different diseases, is like a paradox in Salmonella biology. Repeat is one of the probes that can explain such differences. Here, a comparative genomics approach is followed to identify and characterize repeats that might play role in adaptation and pathogenesis. Repeats are non-randomly distributed in the genomes except few typhoid causing strains. Perfect long repeats are rare compare to polymorphic ones and both are statistically consistent. Significant differences in repeat densities in stress related genes manifest its probable participation in survival and virulence. 573 and 1053 repeat loci have been identified which are exclusively associated with stress and virulent genes respectively. In Salmonella Typhi, an octameric VNTR locus is found in between acrD and yffB genes having more than 25 perfect copies across Salmonella Typhi but possesses only single copy in other serovars. This repeat can be used as a diagnostic probe for typhoid.

Microsatellite marker development by partial sequencing of the sour passion fruit genome (Passiflora edulis Sims)

BackgroundThe Passiflora genus comprises hundreds of wild and cultivated species of passion fruit used for food, industrial, ornamental and medicinal purposes. Efforts to develop genomic tools for genetic analysis of P. edulis, the most important commercial Passiflora species, are still incipient. In spite of many recognized applications of microsatellite markers in genetics and breeding, their availability for passion fruit research remains restricted. Microsatellite markers in P. edulis are usually limited in number, show reduced polymorphism, and are mostly based on compound or imperfect repeats. Furthermore, they are confined to only a few Passiflora species. We describe the use of NGS technology to partially assemble the P. edulis genome in order to develop hundreds of new microsatellite markers.ResultsA total of 14.11 Gbp of Illumina paired-end sequence reads were analyzed to detect simple sequence repeat sites in the sour passion fruit genome. A sample of 1300 contigs containing perfect repeat microsatellite sequences was selected for PCR primer development. Panels of di- and tri-nucleotide repeat markers were then tested in P. edulis germplasm accessions for validation. DNA polymorphism was detected in 74% of the markers (PIC = 0.16 to 0.77; number of alleles/locus = 2 to 7). A core panel of highly polymorphic markers (PIC = 0.46 to 0.77) was used to cross-amplify PCR products in 79 species of Passiflora (including P. edulis), belonging to four subgenera (Astrophea, Decaloba, Distephana and Passiflora). Approximately 71% of the marker/species combinations resulted in positive amplicons in all species tested. DNA polymorphism was detected in germplasm accessions of six closely related Passiflora species (P. edulis, P. alata, P. maliformis, P. nitida, P. quadrangularis and P. setacea) and the data used for accession discrimination and species assignment.ConclusionA database of P. edulis DNA sequences obtained by NGS technology was examined to identify microsatellite repeats in the sour passion fruit genome. Markers were submitted to evaluation using accessions of cultivated and wild Passiflora species. The new microsatellite markers detected high levels of DNA polymorphism in sour passion fruit and can potentially be used in genetic analysis of P. edulis and other Passiflora species.

Phylogenetic studies and comparative chloroplast genome analyses elucidate the basal position of halophyte Nitraria sibirica (Nitrariaceae) in the Sapindales

Nitraria sibirica is a halophyte and belongs to the family Nitrariaceae. The chloroplast genome of Nitraria sibirica (159,466 bp) has a quadripartite structure, which consists of a large single-copy (87,914 bp) region, a small single-copy (18,316 bp) region, and a pair of inverted repeats (26,618 bp). Sequencing analyses indicate that the chloroplast genome contains 113 distinct genes, including 79 peptide-coding genes, 30 transfer RNA genes, and 4 ribosomal RNA genes. We also identified 105 perfect simple sequence repeats, 12 most divergent non-coding regions, and 6 most divergent coding regions when compared to the chloroplast genomes of the Sapindales plants. Phylogenetic analyses using the concatenated amino acid sequences of 58 protein-coding genes from 48 species suggest that the ‘basal’ position of Nitraria sibirica belongs to the Sapindales clade. We also found that the inverted repeat expansion resulted in a duplication of rps19 in Nitraria sibirica when comparing its chloroplast genome structure with Theobroma cacao, Vitis vinifera, Eucalyptus erythrocorys and Arabidopsis thaliana. The duplication of rps19 gene was consistent with that in Zanthoxylum piperitum, Azadirachta indica, Sapindus mukorossi and Citrus sinensis, all of which belong to the order Sapindales, but different from most Rosids plants. In summary, the analyses of Nitraria sibirica chloroplast genome not only provide insights into comparative genome analysis, but also pave the way for a better understanding of the phylogenetic relationships within the Sapindales.

The genome sequence of Condylorrhiza vestigialis NPV, a novel baculovirus for the control of the Alamo moth on Populus spp. in Brazil.

Condylorrhiza vestigialis (Lepidoptera: Cambridae), commonly known as the Brazilian poplar moth or Alamo moth, is a serious defoliating pest of poplar, a crop of great economic importance for the production of wood, fiber, biofuel and other biomaterials as well as its significant ecological and environmental value. The complete genome sequence of a new alphabaculovirus isolated from C. vestigialis was determined and analyzed. Condylorrhiza vestigialis nucleopolyhedrovirus (CoveNPV) has a circular double-stranded DNA genome of 125,767bp with a GC content of 42.9%. One hundred and thirty-eight putative open reading frames were identified and annotated in the CoveNPV genome, including 38 core genes and 9 bros. Four homologous regions (hrs), a feature common to most baculoviruses, and 19 perfect and imperfect direct repeats (drs) were found. Phylogenetic analysis confirmed that CoveNPV is a Group I Alphabaculovirus and is most closely related to Anticarsia gemmatalis multiple nucleopolyhedrovirus (AgMNPV) and Choristoneura fumiferana DEF multiple nucleopolyhedrovirus CfDEFMNPV. The gp37 gene was not detected in the CoveNPV genome, although this gene is found in many NPVs. Two other common NPV genes, chitinase (v-chiA) and cathepsin (v-cath), that are responsible for host insect liquefaction and melanization, were also absent, where phylogenetic analysis suggests that the loss these genes occurred in the common ancestor of AgMNPV, CfDEFMNPV and CoveNPV, with subsequent reacquisition of these genes by CfDEFMNPV. The molecular biology and genetics of CoveNPV was formerly very little known and our expectation is that the findings presented here should accelerate research on this baculovirus, which will facilitate the use of CoveNPV in integrated pest management programs in Poplar crops.

Characterization by next-generation sequencing of 24 new microsatellite loci for the barred sand-bass, Paralabrax nebulifer (Girard, 1854), from the Baja California Peninsula, Mexico

We characterized a set of new hypervariable microsatellite loci for the barred sand-bass (Paralabrax nebulifer), a marine fish that supports important recreational and artisanal fisheries in California, USA and the west coast of the Baja California Peninsula, Mexico. We performed a shotgun genome sequencing with the 454 XL titanium chemistry and used bioinformatics to search for microsatellite loci with perfect repeats. We selected 40 primer pairs that were synthesized and genotyped in an ABI PRISM 3730XL DNA sequencer in 32 individuals from San Juanico, Baja California Sur. We estimated levels of genetic diversity, deviations from linkage and Hardy–Weinberg equilibrium, the frequency of null alleles and the probability of individual identity for the new markers. We successfully scored 24 microsatellite loci (13 tetranucleotides and 11 dinucleotides). The average number of alleles per locus was 12.5 (range 4–23). The average observed and expected heterozygosities were 0.779 (range 0.313–0.969) and 0.774 (range 0.350–0.939), respectively. We detected significant linkage disequilibrium in two pairs of loci. Genotype frequencies at seven loci showed significant deviations from the expectations of Hardy–Weinberg equilibrium and had estimated null allele frequencies ≥10%. The probability of individual identity for the new loci was 8.5−36. The new markers will be useful for investigating patterns of fine-scale genetic structure and diversity to estimate larval dispersal and assess metapopulation dynamics, information necessary for the sustainable management of P. nebulifer fisheries at the west coast of the Baja California Peninsula.

Transposition of the bamboo Mariner-like element Ppmar1 in yeast

The moso bamboo genome contains the two structurally intact and thus potentially functional mariner-like elements Ppmar1 and Ppmar2. Both elements contain perfect terminal inverted repeats (TIRs) and a full-length intact transposase gene. Here we investigated whether Ppmar1 is functional in yeast (Saccharomyces cerevisiae). We have designed a two-component system consisting of a transposase expression cassette and a non-autonomous transposon on two separate plasmids. We demonstrate that the Ppmar1 transposase Pptpase1 catalyses excision of the non-autonomous Ppmar1NA element from the plasmid and reintegration at TA dinucleotide sequences in the yeast chromosomes. In addition, we generated 14 hyperactive Ppmar1 transposase variants by systematic single amino acid substitutions. The most active transposase variant, S171A, induces 10-fold more frequent Ppmar1NA excisions in yeast than the wild type transposase. The Ppmar1 transposon is a promising tool for insertion mutagenesis in moso bamboo and may be used in other plants as an alternative to the established transposon tagging systems.

Two active bamboo mariner-like transposable elements (Ppmar1 and Ppmar2) identified as the transposon-based genetic tools for mutagenesis

Bamboo is one of the most important non-timber forest species in the world, but their molecular breeding lags far behind in contrast to other economic plants. Regarding the difficulties of hybridization and gene modification, the transposon-based insertional mutagenesis might be an alternative, feasible way for molecular breeding of bamboo. A systematic search for potential active transposons identified two full-length mariner-like elements (MLEs) (Ppmar1 and Ppmar2) from moso bamboo in the previous study. Both MLEs contain perfect terminal inverted repeats (TIRs) and a full-length intact transposase. Two transposases contain intact DNA-binding motifs and a DD39D catalytic domain which indicates that Ppmar1 and Ppmar2 are likely active. Here, we deployed a heterologous transposition system of Arabidopsis thaliana to study the transposition activity of Ppmar1 and Ppmar2. The results show that both MLEs could transpose in A. thaliana. Excisions of Ppmar1 and Ppmar2 are usually unperfect as they leave 1–4 bp in excision sites. The reinsertions of both Ppmar1 and Ppmar2 occur at TA dinucleotides and prefer to insert into the TA-rich regions. The insertion sites are dispersed and non-linked. Two active bamboo transposons identified here not only could be applied to construction of the bamboo mutant libraries but also would provide another choice for other plant transposon-based gene tagging.

A Genome-Wide Survey of the Microsatellite Content of the Globe Artichoke Genome and the Development of a Web-Based Database.

The recently acquired genome sequence of globe artichoke (Cynara cardunculus var. scolymus) has been used to catalog the genome’s content of simple sequence repeat (SSR) markers. More than 177,000 perfect SSRs were revealed, equivalent to an overall density across the genome of 244.5 SSRs/Mbp, but some 224,000 imperfect SSRs were also identified. About 21% of these SSRs were complex (two stretches of repeats separated by <100 nt). Some 73% of the SSRs were composed of dinucleotide motifs. The SSRs were categorized for the numbers of repeats present, their overall length and were allocated to their linkage group. A total of 4,761 perfect and 6,583 imperfect SSRs were present in 3,781 genes (14.11% of the total), corresponding to an overall density across the gene space of 32,5 and 44,9 SSRs/Mbp for perfect and imperfect motifs, respectively. A putative function has been assigned, using the gene ontology approach, to the set of genes harboring at least one SSR. The same search parameters were applied to reveal the SSR content of 14 other plant species for which genome sequence is available. Certain species-specific SSR motifs were identified, along with a hexa-nucleotide motif shared only with the other two Compositae species (sunflower (Helianthus annuus) and horseweed (Conyza canadensis)) included in the study. Finally, a database, called “Cynara cardunculus MicroSatellite DataBase” (CyMSatDB) was developed to provide a searchable interface to the SSR data. CyMSatDB facilitates the retrieval of SSR markers, as well as suggested forward and reverse primers, on the basis of genomic location, genomic vs genic context, perfect vs imperfect repeat, motif type, motif sequence and repeat number. The SSR markers were validated via an in silico based PCR analysis adopting two available assembled transcriptomes, derived from contrasting globe artichoke accessions, as templates.

Population study and mutation analysis for 28 short tandem repeat loci in southwest Chinese Han population

Short tandem repeat (STR) system is the most widely used genetic markers in modem forensic practice. Because of the relatively unstable molecular structure, STRs show a high mutation rate. In the current study, we report 169 mutation events of 13 CODIS and 15 non-CODIS STR loci that were found in 5569 cases of trios and duos paternity test. Our result indicated that locus-specific mutation rate varied among different populations, geometric means of the longest run of perfect repeats (LRPR) and heterozygosity. Along with previous published data, a forensic dataset for allele frequencies and locus-specific mutation rates of 13 CODIS and 15 non-CODIS STR loci from southwest Chinese Han population has been established. The mutation rate data have important implications in interpreting forensic individual identification and paternity testing.

Novel microsatellite markers for Distylium lepidotum (Hamamelidaceae) endemic to the Ogasawara Islands.

BackgroundDistylium lepidotum is a small tree endemic to the Ogasawara Islands located in the northwestern Pacific Ocean. This species is a sole food for an endemic locust, Boninoxya anijimensis. Here, we developed microsatellite markers to investigate genetic diversity and genetic structure and to avoid a genetic disturbance after transplantation to restore the Ogasawara Islands ecosystem.ResultsMicrosatellite markers with perfect dinucleotide repeats were developed using the next-generation sequencing Illumina MiSeq Desktop Sequencer. Thirty-two primer pairs were characterized in two D. lepidotum populations on Chichijima and Hahajima Islands of the Ogasawara Islands. The number of alleles for the markers ranged from three to 23 per locus in the two populations. Expected heterozygosity per locus in each population ranged from 0.156 to 0.940 and 0.368 to 0.845, respectively.ConclusionsThese microsatellite markers will be useful for future population genetics studies of D. lepidotum and provide a basis for conservation management of the Ogasawara Islands.

Microsatellites mining in date palm (Phoenix dactylifera L.) and their cross transferability across Arecaceae family

Phoenix dactylifera L. (date palm) is the first publicly available nuclear genome sequence under Arecaceae family and indeed under the entire order Arecales. These genomic sequences were mined for microsatellites which could be used for marker assisted selection for important traits. We have developed an integrated, configurable and time effective microsatellite mining and annotation pipeline for this purpose. Microsatellite survey of date palm whole genome shotgun sequences using the developed pipeline detected a total of 166,760 perfect repeats with an average of one SSR per 2.2kb. Putative functions of these SSR-containing sequences within the proximity of genic regions were predicted. Primers targeting the functionally important SSR regions were designed and a set of eight SSR primers synthesized and were validated in vitro across five members of the Arecaceae family (coconut, arecanut, oilpalm, date palm and palmyrah palm). We have found that 28.6% of the SSRs are common among the members of arecaceae family. Those SSRs could be used in molecular marker analysis of less studied palm family members like arecanut and palmyrah palm.

Mutational analysis of 33 autosomal short tandem repeat (STR) loci in southwest Chinese Han population based on trio parentage testing

Mutation rates and 95% CI of 33 short tandem repeat (STR) loci (D1S2142, D2S1338, D2S441, D3S1358, D3S1754, D5S818, D6S1043, D7S3048, D7S820, D8S1132, D8S1179, D10S1248, D11S2368, D12S391, D13S1492, D13S317, D13S325, D14S306, D15S659, D16S539, D18S1364, D18S51, D19S433, D20S161, D21S11, D22GATA198B05, CSF1PO, FGA, Penta D, Penta E, TH01, TPOX, and vWA) were investigated through more than 424,000 parent-child meiotic transfers obtained from 10636 trios parentage testing cases in southwest Chinese Han population. Overall, 297, including 292 single-step, 4 double-step and 1 triple-step mutation events were observed. The average mutation rate was 0.70×10−3. Most of the locus-specific mutation rates (varied from 0.20×10−3 to 1.96×10−3) were lower than the other datasets (p<0.05). Mutations of 7 loci are reported for the first time. Mutation rates varied with population from different ethnicities and geographical regions. There was no significant difference between mutation expansion and contraction (∼1.04:1). Paternal origin mutations occurred more frequently than maternal origin ones (∼5.02:1). In addition, mutation rates indicated positive correlation with the expected heterozygosity (He) and geometric mean of longest run of perfect repeats (LRPR), respectively. Short alleles showed a trend toward mutation gain while long alleles trended toward mutation loss. A credible forensic dataset for locus-specific mutation rates of 33 loci has been established based upon strict inclusion criteria of large-sized parents/child-trio cases.

Repeat sequence analysis of Mycobacterium tuberculosis

Tuberculosis is a major cause of human death around the world and is caused by various strains of mycobacteria, usually Mycobacterium tuberculosis. The bacterium infects 1.8 billion people yearly which equals one-third of the world population. Pathogenic bacterial genomes contain many perfect, imperfect and approximate tandem repeats that can serve as marker for genotyping these pathogens. Tandem repeats are generated by duplications during successive generations which changes genome structures, thereby providing diversity and improving the fitness of the pathogen during infection. In the present study, we have retrieved reference genomes of 30 Mycobacterium tuberculosis strains and used Tandem Repeats Finder tool for the prediction of genome-wide repeats. The genomes of publicly available Mycobacterium tuberculosis strains varied in size from 4.38 MB to 4.42 MB. After trimming out low quality tandem repeats, we found the strain M. tuberculosis CCDC5079 to have the maximum tandem repeats density number (DN) while density length (DL) was found to be maximum in M. tuberculosis KIT87190.We found the lowest repeat density number and length in M. tuberculosis Erdman. This study would help in mapping and identification of pathogenic bacterial species and to understand the importance of tandem repeats in their evolution.

Identification and characterization of microsatellite markers in Penstemon scariosus (Plantaginaceae).

Premise of the study:Penstemon scariosus var. albifluvis (Plantaginaceae) has been proposed to be federally listed as threatened due to its unique, geologically oil-rich habitat. Developing simple sequence repeat (SSR) markers to study its genetic diversity would be most useful.Methods and Results:Using genomic reduction in combination with next-generation sequencing, we identified SSR motifs with five to 15 perfect repeats in 1067 P. scariosus contigs. After multiple qualifying tests, 16 SSRs were selected for their robust polymorphic reliability across 12 taxa with as high as 21 alleles in a given taxon. With the exception of two monomorphic loci, the observed and expected heterozygosity values ranged from 0.083 to 1.000 and 0.398 to 0.920, respectively.Conclusions:These microsatellite markers will directly aid in studies of the genetic diversity and relatedness of P. scariosus, P. comarrhenus, P. compactus, P. cyananthus var. cyananthus, P. fremontii var. fremontii, P. fremontii var. glabrescens, P. gibbensii, P. strictus, and P. subglaber.