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Eldar Markov
Eldar Markov

Lightcycler Software 4.1 |WORK| Download 17


Opening new horizons in the field of data analysis, the LightCycler Software maintains and expands the analysis modules for absolute quantification and melting curve analysis. Additionally, the new software integrates a dedicated relative quantification module for mono- and dual-color experiments. A module for genotyping by melting curve analysis is also part of the package. Most analysis modules automate the analysis of internal controls.




lightcycler software 4.1 download 17


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(A) Multiple alignment of selected fish IL-17A/F2 proteins with representative mammalian IL-17A and IL-17F molecules. Conserved residues are shaded using BOXSHADE (v3.21), with Cys residues indicated by black characters on a gray background. The predicted signal peptide (SP) is indicated by a black solid double-headed arrow above the alignment. The four cysteine residues that form two potential disulfide bonds are also indicated. Identical residues are identified by stars, whereas similar residues are identified by single dots. (B) Phylogenetic tree showing the relationship of the trout IL-17A/F2 amino acid sequence with those of other known IL-17 family members. The amino acid sequences were aligned using CLUSTAL W, and the tree was constructed by the N-J method supported with 10,000 bootstrap replications using MEGA 4.1 software. The trout IL-17A/F2 protein is in bold and underlined. Accession numbers of the IL-17 family members used are as follows: for IL-17A, human, "type":"entrez-protein","attrs":"text":"NP_002181","term_id":"4504651"NP_002181; cow, "type":"entrez-protein","attrs":"text":"NP_001008412","term_id":"56605852"NP_001008412; mouse, "type":"entrez-protein","attrs":"text":"NP_034682","term_id":"6754324"NP_034682; rat, "type":"entrez-protein","attrs":"text":"Q61453","term_id":"2498483"Q61453; and chicken, "type":"entrez-protein","attrs":"text":"CAO79600","term_id":"194326351"CAO79600; for IL-17F, human, "type":"entrez-protein","attrs":"text":"NP_443104","term_id":"16418375"NP_443104; cow, "type":"entrez-protein","attrs":"text":"NP_001179011","term_id":"300797226"NP_001179011; mouse, "type":"entrez-protein","attrs":"text":"NP_665855","term_id":"119360346"NP_665855; rat, "type":"entrez-protein","attrs":"text":"NP_001015011","term_id":"845634515"NP_001015011; and chicken, "type":"entrez-protein","attrs":"text":"NP_989791","term_id":"2103987647"NP_989791; for IL-17A/F2, zebrafish, NP_00101863; fugu, "type":"entrez-protein","attrs":"text":"BAI82579","term_id":"291202720"BAI82579; and medaka, "type":"entrez-protein","attrs":"text":"NP_001191713","term_id":"325652194"NP_001191713; for IL-17A/F1, fugu, "type":"entrez-protein","attrs":"text":"BAI82578","term_id":"291202718"BAI82578; medaka, "type":"entrez-protein","attrs":"text":"NP_001191714","term_id":"325652196"NP_001191714; and zebrafish, NP_00101862; for IL-17A/F3, zebrafish, NP_00101862; medaka, "type":"entrez-protein","attrs":"text":"NP_001191715","term_id":"325652158"NP_001191715; and fugu, "type":"entrez-protein","attrs":"text":"BAI82580","term_id":"291202722"BAI82580; for IL-17C, C1 trout, "type":"entrez-protein","attrs":"text":"CAW30794","term_id":"291196115"CAW30794; C2 trout, "type":"entrez-protein","attrs":"text":"NP_001171959","term_id":"297206847"NP_001171959; zebrafish, "type":"entrez-protein","attrs":"text":"NP_001018624","term_id":"66472828"NP_001018624; fugu, "type":"entrez-protein","attrs":"text":"BAI82581","term_id":"294774540"BAI82581; medaka, "type":"entrez-protein","attrs":"text":"NP_001191723","term_id":"325652180"NP_001191723; human, "type":"entrez-protein","attrs":"text":"NP_037410","term_id":"7019435"NP_037410; cow, "type":"entrez-protein","attrs":"text":"DAA20250","term_id":"296478135"DAA20250; mouse, "type":"entrez-protein","attrs":"text":"NP_665833","term_id":"1215189884"NP_665833; and rat, NP_0011780; for IL-17E, human, "type":"entrez-protein","attrs":"text":"AAQ89484","term_id":"37183370"AAQ89484; mouse, "type":"entrez-protein","attrs":"text":"NP_542767","term_id":"18141561"NP_542767; cow, "type":"entrez-protein","attrs":"text":"DAA25796","term_id":"296483681"DAA25796; dog, XP_537375; and horse, XP_001918360; for IL-17B, mouse, "type":"entrez-protein","attrs":"text":"NP_062381","term_id":"9507247"NP_062381; rat, "type":"entrez-protein","attrs":"text":"NP_446241","term_id":"281371353"NP_446241; human, "type":"entrez-protein","attrs":"text":"CAG33473","term_id":"48146501"CAG33473; chicken, XP_425192; and xenopus, "type":"entrez-protein","attrs":"text":"AAH75405","term_id":"49522392"AAH75405; and for IL-17D, human, "type":"entrez-protein","attrs":"text":"NP_612141","term_id":"19923715"NP_612141; mouse, "type":"entrez-protein","attrs":"text":"NP_665836","term_id":"148277104"NP_665836; cow, "type":"entrez-protein","attrs":"text":"DAA23895","term_id":"296481780"DAA23895; medaka, "type":"entrez-protein","attrs":"text":"NP_001191716","term_id":"325652166"NP_001191716; fugu, "type":"entrez-protein","attrs":"text":"AAI58483","term_id":"165970584"AAI58483; zebrafish, "type":"entrez-protein","attrs":"text":"AAI62897","term_id":"190338167"AAI62897; trout, "type":"entrez-protein","attrs":"text":"CAE45584","term_id":"33945897"CAE45584; and salmon, "type":"entrez-protein","attrs":"text":"NP_001134365","term_id":"213514562"NP_001134365.


(A) Multiple alignment of mammalian and fish IL-17RA protein sequences. Conserved residues are shaded using BOXSHADE (v3.21), with Cys residues indicated by black characters on a gray background. The predicted signal peptide (SP), D1 and D2 domains, transmembrane region (TM), SEFIR domain, and repeats (R) are indicated by black solid double-headed arrows above the alignment. The eight cysteine residues that form four potential disulfide bonds are also indicated. Identical residues are identified by stars, whereas similar residues are identified by single dots. (B) Phylogenetic tree showing the relationship of trout receptors with other known IL-17R family members. The amino acid sequences were aligned using CLUSTAL W, and the tree was constructed by the N-J method supported with 1,000 bootstrap replications using MEGA 4.1 software. The trout IL-17RA protein is in bold and underlined. Accession numbers of the sequences used in the analysis are as follows: for IL-17RA, salmon, "type":"entrez-protein","attrs":"text":"NP_001158836","term_id":"259155188"NP_001158836; zebrafish, XP_001921444; chicken, XP_416389; human, "type":"entrez-protein","attrs":"text":"CAJ86450","term_id":"90403048"CAJ86450; and mouse, "type":"entrez-protein","attrs":"text":"NP_032385","term_id":"6680411"NP_032385; for IL-17RD, zebrafish, "type":"entrez-protein","attrs":"text":"AAI63933","term_id":"190337798"AAI63933; xenopus, "type":"entrez-protein","attrs":"text":"XP_002940076","term_id":"301621474"XP_002940076; chicken, "type":"entrez-protein","attrs":"text":"NP_989846","term_id":"2099389656"NP_989846; human, "type":"entrez-protein","attrs":"text":"NP_060033","term_id":"166063983"NP_060033; and mouse, "type":"entrez-protein","attrs":"text":"NP_602319","term_id":"24025662"NP_602319; for IL-17RB, cow, "type":"entrez-protein","attrs":"text":"AAI33637","term_id":"126920901"AAI33637; human, "type":"entrez-protein","attrs":"text":"NP_061195","term_id":"27477074"NP_061195; mouse, "type":"entrez-protein","attrs":"text":"NP_062529","term_id":"83025064"NP_062529, and rat, "type":"entrez-protein","attrs":"text":"NP_001100760","term_id":"157822013"NP_001100760; for IL-17RC, rat, "type":"entrez-protein","attrs":"text":"NP_001164036","term_id":"281604108"NP_001164036; human, "type":"entrez-protein","attrs":"text":"NP_116121","term_id":"1676319182"NP_116121; mouse, "type":"entrez-protein","attrs":"text":"NP_598920","term_id":"282399145"NP_598920; and cow, "type":"entrez-protein","attrs":"text":"NP_001068646","term_id":"115496380"NP_001068646; and for IL-17RE, rat, "type":"entrez-protein","attrs":"text":"NP_001004091","term_id":"51890226"NP_001004091; mouse, "type":"entrez-protein","attrs":"text":"AAH69861","term_id":"47122706"AAH69861; human, "type":"entrez-protein","attrs":"text":"NP_705613","term_id":"24430204"NP_705613; and cow, XP_001251391.


The population divergence was measured by calculating the fixation index (FST) for all pairs of populations. The online software, COIL (Broad Institute, USA) was used to determine the FST for both populations.


We also wish to underline that whilst the above conclusions are particularly relevant to whole-genome resequencing of organisms from which high quality DNA is routinely available (e.g. human WGS), alternative applications of WGS may benefit from different choices of insert length. For example, applications that entail low-coverage sequencing such as genome-skimming, may benefit from avoiding a wide range of insert sizes and resulting wider range of genome coverage, as this may in some instances lead to consistent drop-out of low-coverage areas, despite a higher average coverage [18, 19]. Overlapping reads not only increase the accuracy of base-calls, but the resulting longer merged reads can aid in read mapping, particularly if ambiguous base calls are present. Applications that may benefit from the deliberate use of overlapping paired end reads include ancient-DNA and forensic sequencing [20]. In addition, the software and analysis strategy chosen should also be considered on a per-application basis prior to choosing a desired insert length (for example see [21]).


A recent publication detected increased insertion /deletion artefacts in libraries prepared with enzymatic fragmentation relative to those prepared by sonication [22]. Although the authors produced a software tool to detect such artefacts, this finding reinforces the reputation of sonication as the gold-standard method of DNA fragmentation, even though some simple steps must also be taken with sonication to avoid introducing sequencing artefacts [23]. Nonetheless, we speculate that our observation that library insert size affects genome coverage and variant detection will also be valid for libraries produced by sonication.


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