BACKGROUND: The program InDeVal was originally developed to help researchers find known regions of insertion/deletion activity (with the exception of isolated single-base indels) in newly determined Poaceae trnL-F sequences and compare them with 533 previously determined sequences. It is supplied with input files designed for this purpose. More broadly, the program is applicable for finding specific target regions (referred to as "variable regions") in DNA sequence. A variable region is any specific sequence fragment of interest, such as an indel region, a codon or codons, or sequence coding for a particular RNA secondary structure. RESULTS: InDeVal input is DNA sequence and a template file (sequence flanking each variable region). Additional files contain the variable regions and user-defined messages about the sequence found within them (e.g., taxa sharing each of the different indel patterns).Variable regions are found by determining the position of flanking sequence (referred to as "conserved regions") using the LPAM (Length-Preserving Alignment Method) algorithm. This algorithm was designed for InDeVal and is described here for the first time. InDeVal output is an interactive display of the analyzed sequence, broken into user-defined units. Once the user is satisfied with the organization of the display, the information can be exported to an annotated text file. CONCLUSIONS: InDeVal can find multiple variable regions simultaneously (28 indel regions in the Poaceae trnL-F files) and display user-selected messages specific to the sequence variants found. InDeVal output is designed to facilitate comparison between the analyzed sequence and previously evaluated sequence. The program's sensitivity to different levels of nucleotide and/or length variation in conserved regions can be adjusted. InDeVal is currently available for Windows in Additional file 1 or from http://www.sci.muni.cz/botany/elzdroje/indeval/.

• MeSH
• DNA rostlinná genetika   MeSH
• inzerční mutageneze genetika   MeSH
• kodon genetika   MeSH
• konzervovaná sekvence genetika   MeSH
• lipnicovité genetika   MeSH
• mutační analýza DNA metody   MeSH
• sekvenční delece genetika   MeSH
• sekvenční seřazení metody   MeSH
• software * MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH
• srovnávací studie MeSH
• Názvy látek
• DNA rostlinná MeSH
• kodon MeSH

BACKGROUND: Polyploidization is considered one of the main mechanisms of plant genome evolution. The presence of multiple copies of the same gene reduces selection pressure and permits sub-functionalization and neo-functionalization leading to plant diversification, adaptation and speciation. In bread wheat, polyploidization and the prevalence of transposable elements resulted in massive gene duplication and movement. As a result, the number of genes which are non-collinear to genomes of related species seems markedly increased in wheat. RESULTS: We used new-generation sequencing (NGS) to generate sequence of a Mb-sized region from wheat chromosome arm 3DS. Sequence assembly of 24 BAC clones resulted in two scaffolds of 1,264,820 and 333,768 bases. The sequence was annotated and compared to the homoeologous region on wheat chromosome 3B and orthologous loci of Brachypodium distachyon and rice. Among 39 coding sequences in the 3DS scaffolds, 32 have a homoeolog on chromosome 3B. In contrast, only fifteen and fourteen orthologs were identified in the corresponding regions in rice and Brachypodium, respectively. Interestingly, five pseudogenes were identified among the non-collinear coding sequences at the 3B locus, while none was found at the 3DS locus. CONCLUSION: Direct comparison of two Mb-sized regions of the B and D genomes of bread wheat revealed similar rates of non-collinear gene insertion in both genomes with a majority of gene duplications occurring before their divergence. Relatively low proportion of pseudogenes was identified among non-collinear coding sequences. Our data suggest that the pseudogenes did not originate from insertion of non-functional copies, but were formed later during the evolution of hexaploid wheat. Some evidence was found for gene erosion along the B genome locus.

• MeSH
• Brachypodium genetika   MeSH
• chromozomy rostlin genetika   MeSH
• DNA rostlinná genetika   MeSH
• duplikace genu MeSH
• fylogeneze MeSH
• genetické lokusy genetika   MeSH
• genom rostlinný genetika   MeSH
• inzerční mutageneze MeSH
• kontigové mapování MeSH
• molekulární evoluce * MeSH
• polyploidie MeSH
• pšenice genetika   MeSH
• pseudogeny genetika   MeSH
• rýže (rod) genetika   MeSH
• sekvenční analýza DNA MeSH
• umělé bakteriální chromozomy MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• srovnávací studie MeSH
• Názvy látek
• DNA rostlinná MeSH

BACKGROUND: The insertion sequence elements (IS elements) represent the smallest and the most abundant mobile elements in prokaryotic genomes. It has been shown that they play a significant role in genome organization and evolution. To better understand their function in the host genome, it is desirable to have an effective detection and annotation tool. This need becomes even more crucial when considering rapid-growing genomic and metagenomic data. The existing tools for IS elements detection and annotation are usually based on comparing sequence similarity with a database of known IS families. Thus, they have limited ability to discover distant and putative novel IS elements. RESULTS: In this paper, we present digIS, a software tool based on profile hidden Markov models assembled from catalytic domains of transposases. It shows a very good performance in detecting known IS elements when tested on datasets with manually curated annotation. The main contribution of digIS is in its ability to detect distant and putative novel IS elements while maintaining a moderate level of false positives. In this category it outperforms existing tools, especially when tested on large datasets of archaeal and bacterial genomes. CONCLUSION: We provide digIS, a software tool using a novel approach based on manually curated profile hidden Markov models, which is able to detect distant and putative novel IS elements. Although digIS can find known IS elements as well, we expect it to be used primarily by scientists interested in finding novel IS elements. The tool is available at https://github.com/janka2012/digIS.

• Klíčová slova
• Genome annotation, IS elements, Mobile element, Profile HMM, Prokaryotic genomes,
• MeSH
• genom bakteriální genetika   MeSH
• genomika MeSH
• lidé MeSH
• prokaryotické buňky * MeSH
• software MeSH
• transpozibilní elementy DNA * genetika   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• transpozibilní elementy DNA * MeSH

Paenibacillus larvae and Melissococcus plutonius represent the most threatening bacterial diseases of honeybee (Apis mellifera)-American and European foulbrood, respectively. For efficient control of those diseases, rapid and accurate detection of the pathogens is crucial. Therefore, we developed a novel multiplex PCR method simultaneously detecting both pathogens. To design and optimize multiplex PCR reaction, four strains of P. larvae representing four ERIC genotypes I-IV (strain DSM 7030-ERIC I, DSM 25430-ERIC II, LMG 16252-ERIC III, DSM 3615-ERIC IV) were selected. Those strains were fully sequenced using long-read sequencing (Sequel I, Pacific Biosciences). For P. larvae, the multicopy insertion sequence IS256 identified in all genotypes of P. larvae was selected to provide high sensitivity. M. plutonius was detected by plasmid pMP1 sequence and the virulence verified by following detection of ETX/MTX2 toxin responsible for pore formation in the cell membrane. As an internal control, a gene encoding for major royal jelly protein 1 specific for honeybees was selected. The method was validated on 36 clinical specimens collected from the colonies suffering from American and European foulbrood in the Czech Republic. Based on the results, sensitivity of PCR was calculated to 93.75% and specificity to 100% for P. larvae diagnosed from hive debris and 100% sensitivity and specificity for honeybee workers and larval scales as well as for diseased brood infected by M. plutonius.

• Klíčová slova
• Apis mellifera, Melissococcus plutonius, Paenibacillus larvae, American foulbrood, European foulbrood, Multiplex PCR,
• MeSH
• Enterococcaceae * MeSH
• larva mikrobiologie   MeSH
• multiplexová polymerázová řetězová reakce metody   MeSH
• Paenibacillus larvae * genetika   MeSH
• Paenibacillus * genetika   MeSH
• plazmidy genetika   MeSH
• transpozibilní elementy DNA MeSH
• včely genetika   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• transpozibilní elementy DNA MeSH

Burkholderia cenocepacia can cause serious infections and epidemics in patients with cystic fibrosis (CF). A CF population in the Czech Republic experienced an epidemic outbreak caused by a B. cenocepacia ST-32 strain. The clonality of the isolates was evident by multilocus sequence typing; however, fingerprinting profiles obtained by pulsed-field gel electrophoresis (PFGE) showed substantial band variability. We investigated whether the PFGE pattern diversity resulted from genomic rearrangements mediated by insertion sequences (IS); in addition, we determined whether stressful growth conditions altered the transposition activity of these IS. DNA probes for IS commonly found in B. cenocepacia were designed using the B. cenocepacia J2315 genome. Southern hybridization analysis of ST-32 isolates demonstrated diversity in both the copy number and the insertion site for a homologue of ISBcen20. Movement of the ISBcen20 homologue was detected when the ST-32 isolate CZ1238 was exposed to oxidative stress (growth in the presence of H(2)O(2)). PFGE analysis of CZ1238 derivatives exposed to oxidative stress demonstrated genomic rearrangements. Interestingly, when the closely related B. cenocepacia strain J2315 was exposed to oxidative stress, no movement of ISBcen20 was detected. Since frameshift mutations are present within the transposases of all copies of this IS in J2315, our data suggest that the transposase is inactive. In summary, we have demonstrated for the first time that IS movement can be mediated by oxidative stress and can lead to genomic rearrangements in the CF pathogen B. cenocepacia. These IS movements may alter the PFGE fingerprints of isolates that are clonal by other typing methods.

• MeSH
• Burkholderia klasifikace   genetika   izolace a purifikace   MeSH
• cystická fibróza komplikace   MeSH
• DNA bakterií genetika   MeSH
• DNA fingerprinting metody   MeSH
• epidemický výskyt choroby MeSH
• genetická variace * MeSH
• genotyp MeSH
• infekce bakteriemi rodu Burkholderia epidemiologie   mikrobiologie   MeSH
• lidé MeSH
• molekulární epidemiologie metody   MeSH
• oxidační stres * MeSH
• pulzní gelová elektroforéza metody   MeSH
• rekombinace genetická * MeSH
• shluková analýza MeSH
• techniky typizace bakterií metody   MeSH
• transpozibilní elementy DNA MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Česká republika epidemiologie   MeSH
• Názvy látek
• DNA bakterií MeSH
• transpozibilní elementy DNA MeSH

The VERNALIZATION1 (VRN1) gene encodes a MADS-box transcription factor and plays an important role in the cold-induced transition from the vegetative to reproductive stage. Allelic variability of VRN1 homoeologs has been associated with large differences in flowering time. The aim of this study was to investigate the genetic variability of VRN1 homoeologs (VRN-A1, VRN-B1 and VRN-D1). We performed an in-depth sequence analysis of VRN1 homoeologs in a panel of 105 winter and spring varieties of hexaploid wheat. We describe the novel allele Vrn-B1f with an 836 bp insertion within intron 1 and show its specific expression pattern associated with reduced heading time. We further provide the complete sequence of the Vrn-A1b allele, revealing a 177 bp insertion in intron 1, which is transcribed into an alternative splice variant. Copy number variation (CNV) analysis of VRN1 homoeologs showed that VRN-B1 and VRN-D1 are present in only one copy. The copy number of recessive vrn-A1 ranged from one to four, while that of dominant Vrn-A1 was one or two. Different numbers of Vrn-A1a copies in the spring cultivars Branisovicka IX/49 and Bastion did not significantly affect heading time. We also report on the deletion of secondary structures (G-quadruplex) in promoter sequences of cultivars with more vrn-A1 copies.

• Klíčová slova
• CNV, VRN1, allelic variation, alternative splice variants, next generation sequencing, wheat,
• MeSH
• alely * MeSH
• alternativní sestřih MeSH
• chléb MeSH
• genetická variace * MeSH
• genová dávka * MeSH
• inzerční mutageneze MeSH
• polyploidie * MeSH
• pšenice genetika   MeSH
• represorové proteiny genetika   MeSH
• sekvenční analýza DNA MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• represorové proteiny MeSH

The molecular diagnosis of pertussis and parapertussis syndromes is based on the detection of insertion sequences (IS) 481 and 1001, respectively. However, these IS are also detected in the genomes of various Bordetella species, such that they are not specific for either B. pertussis or B. parapertussis. Therefore, we screened the genome of recently circulating isolates of Bordetella species to compare the prevalence of IS481, IS1001 and, also IS1002 with previously published data and to sequence all IS detected. We also investigated whether the numbers of IS481 and IS1001 copies vary in recently circulating isolates of the different Bordetella species. We used the polymerase chain reaction (PCR) method for screening the genome of circulating isolates and to prepare the fragments for sequencing. We used Southern blotting and quantitative real-time PCR for quantification of the numbers of IS. We found no significant diversity in the sequences of the IS harboured in the genomes of the Bordetella isolates screened, except for a 71-nucleotide deletion from IS1002 in B. bronchiseptica. The IS copy numbers in the genome of recently circulating isolates were similar to those in reference strains. Our results confirm that biological diagnosis targeting the IS481 and IS1001 elements are not specific and detect the species B. pertussis, B. holmesii and B. bronchiseptica (IS481), and B. parapertussis and B. bronchiseptica (IS1001).

• MeSH
• bakteriologické techniky metody   MeSH
• Bordetella genetika   izolace a purifikace   MeSH
• diagnostické techniky molekulární metody   MeSH
• DNA bakterií chemie   genetika   MeSH
• genová dávka MeSH
• lidé MeSH
• pertuse diagnóza   mikrobiologie   MeSH
• polymerázová řetězová reakce MeSH
• sekvenční analýza DNA MeSH
• sekvenční delece MeSH
• senzitivita a specificita MeSH
• Southernův blotting MeSH
• transpozibilní elementy DNA * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• DNA bakterií MeSH
• transpozibilní elementy DNA * MeSH

We herein present a rare case of an EML4-ALK positive patient. A 61-year-old man was diagnosed with locoregional non-small cell lung cancer (NSCLC). No EGFR mutations were detected, and therefore the ALK rearrangement was evaluated using immunohistochemistry (IHC), fluorescence in situ hybridization (FISH) and the reverse transcription PCR (RT-PCR) method for EML4-ALK. All methods showed a positive result and, therefore, the patient was treated with crizotinib with a good therapeutic response. However, a detailed RT-PCR analysis and sequencing revealed an unexpected 138 bp insertion of attractin-like 1 (ATRNL1) gene into the EML4-ALK fusion gene. In our case, the positive therapeutic response suggests that ATRNL1 insertion does not affect EML4-ALK's sensitivity to crizotinib. This case shows great EML4-ALK heterogeneity and also that basic detection methods (IHC, FISH) cannot fully specify ALK rearrangement but in many cases a full specification seems to be important for an effective TKI indication, and sequencing ALK variants might contribute to optimized patient selection.

• Klíčová slova
• ATRNL1, Crizotinib, EML4-ALK, Lung cancer, NSCLC, Targeted therapy,
• MeSH
• fúzní onkogenní proteiny chemie   genetika   MeSH
• inhibitory proteinkinas terapeutické užití   MeSH
• inzerční mutageneze * MeSH
• krizotinib MeSH
• lidé středního věku MeSH
• lidé MeSH
• molekulární sekvence - údaje MeSH
• nádory plic diagnóza   farmakoterapie   genetika   MeSH
• nemalobuněčný karcinom plic diagnóza   farmakoterapie   genetika   MeSH
• pyrazoly terapeutické užití   MeSH
• pyridiny terapeutické užití   MeSH
• rentgendiagnostika hrudníku MeSH
• sekvence nukleotidů MeSH
• sekvenční analýza DNA MeSH
• staging nádorů MeSH
• výsledek terapie MeSH
• Check Tag
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH
• práce podpořená grantem MeSH
• Názvy látek
• EML4-ALK fusion protein, human MeSH Prohlížeč
• fúzní onkogenní proteiny MeSH
• inhibitory proteinkinas MeSH
• krizotinib MeSH
• pyrazoly MeSH
• pyridiny MeSH

Renal hypouricemia is a heterogeneous inherited disorder characterized by impaired uric acid handling in the renal tubules. Patients are usually asymptomatic; however, some may experience urolithiasis and/or acute kidney injury. Most of the described patients (compound heterozygous and/or homozygous) are Japanese with mutations in the SLC22A12 gene (OMIM #220150). Four patients with renal hypouricemia caused by heterozygous defects and two families with homozygous mutations in the SLC2A9 gene have been recently described (OMIM #612076). We describe the clinical history, biochemical and molecular genetics findings of a Czech family with renal hypouricemia. The concentration of serum uric acid in the proband (16-year-old Czech girl with unrelated parents) was 0.17 ± 0.05 mg/dl and expressed as an increase in the fractional excretion of uric acid (194 ± 99%). The sequencing analysis of the coding region of uric acid transporters SLC22A12, SLC2A9, SLC17A3, ABCC4 and ABCG2, was performed. Analysis of genomic DNA revealed novel one nucleotide homozygote insertion in exon 3 in the SLC2A9 gene in proband and her brother resulting in a truncated protein (p.Ile118HisfsX27). No sequence variants in other candidate uric acid transporter were found. Homozygous loss-of-function mutations cause massive renal hypouricemia via total loss of uric acid absorption; however, they do not necessarily lead to nephrolithiasis and acute kidney injury. In contrast to previously reported heterozygous patients with renal hypouricemia type 2, we did not find even slight hypouricemia and found no decrease in the FE-UA of the heterozygous parents of the reported siblings.

• MeSH
• genetické asociační studie MeSH
• hodnoty glomerulární filtrace MeSH
• homozygot MeSH
• inzerční mutageneze * MeSH
• kyselina močová krev   MeSH
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• močové kameny diagnóza   genetika   MeSH
• proteiny usnadňující transport glukosy genetika   MeSH
• rodokmen MeSH
• sekvence nukleotidů MeSH
• sekvenční analýza DNA MeSH
• studie případů a kontrol MeSH
• vrozené poruchy tubulárního transportu diagnóza   genetika   MeSH
• Check Tag
• lidé MeSH
• mladiství MeSH
• mladý dospělý MeSH
• mužské pohlaví MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• kazuistiky MeSH
• práce podpořená grantem MeSH
• Názvy látek
• kyselina močová MeSH
• proteiny usnadňující transport glukosy MeSH
• SLC2A9 protein, human MeSH Prohlížeč

Insect silk genes attract attention by their precise territorial and developmental regulations and extremely high expression rates. Our present investigations demonstrated that the P25 silk gene of Galleria mellonella is down-regulated by ecdysteroid hormones. The gene was identified within 5217 nucleotides (nt) of two genomic clones. In contrast to other silk genes, Galleria P25 lacks the canonical TATA box. Transcription is initiated within a region of three nucleotides that lie at the end of a capsite initiator sequence ACAGT and about 90 nt downstream from a CAAT box. A stretch of 32 nt with a core sequence CTTTT was detected in the 5' region of Galleria P25 as well as in the presumptive regulatory regions of all other silk genes that are expressed in the posterior silk gland. However, consensus sequences reported for the regulatory regions of Bombyx silk genes are not obvious in Galleria P25. The coding sequence of this gene included 654 nt, is interrupted by 4 introns, and ends in position +3369; a potential polyadenylation signal starts at +4382. The gene contains 3 copies of a short interspersed nuclear element (SINE), which are located in the upstream region (-833 to -579) and in the first (+542 to +840) and second (+2259 to +2556) introns. The repeat, which was named Gm1, occurs in some other Galleria genes and exhibits homology to Bm1 SINE of the silkworm and to a similar element of a spider. Another insertion of at least 150 nt and with loosely defined borders is present in the 3' untranslated region (UTR) of Galleria P25. It includes a box (+3453 to +3552) of 99 nt that is tentatively called Lep1 because it was disclosed also in some other Lepidoptera. Lep1 seems to represent the core region of insertion elements that occur in the genomes of lepidopteran insects in various species specific and region specific modifications.

• MeSH
• bourec genetika   MeSH
• genetická transkripce MeSH
• genom MeSH
• glykoproteiny biosyntéza   chemie   genetika   MeSH
• hmyzí geny * MeSH
• hmyzí proteiny MeSH
• introny MeSH
• konsenzuální sekvence MeSH
• Lepidoptera genetika   metabolismus   MeSH
• molekulární sekvence - údaje MeSH
• regulační oblasti nukleových kyselin * MeSH
• restrikční mapování MeSH
• sekvence aminokyselin MeSH
• sekvence nukleotidů MeSH
• sekvenční homologie nukleových kyselin MeSH
• sekvenční seřazení MeSH
• transpozibilní elementy DNA * MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• glykoproteiny MeSH
• hmyzí proteiny MeSH
• P25 protein, Galleria mellonella MeSH Prohlížeč
• transpozibilní elementy DNA * MeSH