DOCSLIB.ORG

Mouse Uqcr11 Conditional Knockout Project (CRISPR/Cas9)

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Mouse Uqcr11 Conditional Knockout Project (CRISPR/Cas9) https://www.alphaknockout.com Mouse Uqcr11 Conditional Knockout Project (CRISPR/Cas9) Objective: To create a Uqcr11 conditional knockout Mouse model (C57BL/6J) by CRISPR/Cas-mediated genome engineering. Strategy summary: The Uqcr11 gene (NCBI Reference Sequence: NM_025650 ; Ensembl: ENSMUSG00000020163 ) is located on Mouse chromosome 10. 3 exons are identified, with the ATG start codon in exon 1 and the TAG stop codon in exon 2 (Transcript: ENSMUST00000020372). Exon 2 will be selected as conditional knockout region (cKO region). Deletion of this region should result in the loss of function of the Mouse Uqcr11 gene. To engineer the targeting vector, homologous arms and cKO region will be generated by PCR using BAC clone RP23-132N8 as template. Cas9, gRNA and targeting vector will be co-injected into fertilized eggs for cKO Mouse production. The pups will be genotyped by PCR followed by sequencing analysis. Note: Exon 2 starts from about 30.36% of the coding region. The knockout of Exon 2 will result in frameshift of the gene. The size of intron 1 for 5'-loxP site insertion: 2409 bp, and the size of intron 2 for 3'-loxP site insertion: 980 bp. The size of effective cKO region: ~618 bp. The cKO region does not have any other known gene. Page 1 of 8 https://www.alphaknockout.com Overview of the Targeting Strategy Wildtype allele gRNA region 5' gRNA region 3' 1 2 3 Targeting vector Targeted allele Constitutive KO allele (After Cre recombination) Legends Exon of mouse Uqcr11 Homology arm cKO region loxP site Page 2 of 8 https://www.alphaknockout.com Overview of the Dot Plot Window size: 10 bp Forward Reverse Complement Sequence 12 Note: The sequence of homologous arms and cKO region is aligned with itself to determine if there are tandem repeats. No significant tandem repeat is found in the dot plot matrix. So this region is suitable for PCR screening or sequencing analysis. Overview of the GC Content Distribution Window size: 300 bp Sequence 12 Summary: Full Length(7118bp) | A(23.48% 1671) | C(25.26% 1798) | T(25.5% 1815) | G(25.77% 1834) Note: The sequence of homologous arms and cKO region is analyzed to determine the GC content. No significant high GC-content region is found. So this region is suitable for PCR screening or sequencing analysis. Page 3 of 8 https://www.alphaknockout.com BLAT Search Results (up) QUERY SCORE START END QSIZE IDENTITY CHROM STRAND START END SPAN ----------------------------------------------------------------------------------------------- browser details YourSeq 3000 1 3000 3000 100.0% chr10 - 80404537 80407536 3000 browser details YourSeq 95 742 842 3000 97.1% chr11 - 102492559 102492659 101 browser details YourSeq 90 2262 2972 3000 97.0% chr3 + 104574355 104687771 113417 browser details YourSeq 85 739 842 3000 91.3% chr11 + 52405259 52405365 107 browser details YourSeq 71 772 842 3000 100.0% chr1 - 166431825 166431895 71 browser details YourSeq 59 1255 1370 3000 77.8% chr2 - 163853065 163853179 115 browser details YourSeq 53 2930 2988 3000 96.6% chr19 + 28711235 28711324 90 browser details YourSeq 52 2122 2292 3000 88.1% chr11 - 77201215 77201715 501 browser details YourSeq 51 2934 3000 3000 93.4% chr1 - 15640058 15640152 95 browser details YourSeq 46 2238 2333 3000 87.1% chr11 - 100834558 100834670 113 browser details YourSeq 44 2933 2980 3000 98.0% chr7 - 15960282 15960331 50 browser details YourSeq 44 2930 2981 3000 94.2% chr10 - 44317932 44317985 54 browser details YourSeq 43 2239 2293 3000 89.1% chr16 - 25382599 25382653 55 browser details YourSeq 43 2242 2335 3000 89.1% chr8 + 95851219 95851335 117 browser details YourSeq 43 2947 3000 3000 94.0% chr10 + 5886474 5886554 81 browser details YourSeq 42 2934 2981 3000 95.8% chr11 - 47656823 47656872 50 browser details YourSeq 42 2931 2980 3000 93.9% chr13 + 38685355 38685406 52 browser details YourSeq 41 2237 2334 3000 90.2% chr12 + 109702490 109702593 104 browser details YourSeq 41 2942 2988 3000 95.8% chr1 + 127974360 127974424 65 browser details YourSeq 40 2934 2981 3000 93.7% chr10 - 80203238 80203287 50 Note: The 3000 bp section upstream of Exon 2 is BLAT searched against the genome. No significant similarity is found. BLAT Search Results (down) QUERY SCORE START END QSIZE IDENTITY CHROM STRAND START END SPAN ----------------------------------------------------------------------------------------------- browser details YourSeq 3000 1 3000 3000 100.0% chr10 - 80400919 80403918 3000 browser details YourSeq 229 2201 2519 3000 91.7% chr10 - 121319708 121320480 773 browser details YourSeq 227 2201 2517 3000 87.7% chr12 - 111399645 111399962 318 browser details YourSeq 225 2201 2517 3000 88.2% chr10 - 63180567 63180884 318 browser details YourSeq 225 2205 2517 3000 87.9% chr14 + 65160237 65160550 314 browser details YourSeq 224 2200 2519 3000 89.6% chr10 - 9237736 9238060 325 browser details YourSeq 223 2201 2510 3000 88.9% chr5 + 92984283 92984592 310 browser details YourSeq 221 2201 2512 3000 87.2% chr13 - 46037730 46038042 313 browser details YourSeq 221 2202 2517 3000 89.9% chr13 + 101828777 101829097 321 browser details YourSeq 220 2201 2517 3000 87.0% chr5 - 103217436 103217752 317 browser details YourSeq 220 2196 2517 3000 88.6% chr17 - 29540902 29541227 326 browser details YourSeq 218 2212 2518 3000 89.5% chr9 + 45214736 45215065 330 browser details YourSeq 217 2212 2522 3000 88.1% chr9 - 97189308 97189620 313 browser details YourSeq 217 2204 2518 3000 88.2% chr10 - 105856157 105856472 316 browser details YourSeq 217 2201 2512 3000 90.2% chr1 - 157091528 157109185 17658 browser details YourSeq 215 2212 2518 3000 89.1% chr7 - 82210930 82211238 309 browser details YourSeq 215 2201 2518 3000 90.7% chr1 - 152748125 152748445 321 browser details YourSeq 215 2201 2525 3000 86.7% chr1 + 161738337 161738662 326 browser details YourSeq 215 2204 2512 3000 88.3% chr1 + 128530720 128531260 541 browser details YourSeq 214 2203 2519 3000 87.7% chr7 - 139142554 139142873 320 Note: The 3000 bp section downstream of Exon 2 is BLAT searched against the genome. No significant similarity is found. Page 4 of 8 https://www.alphaknockout.com Gene and protein information: Uqcr11 ubiquinol-cytochrome c reductase, complex III subunit XI [ Mus musculus (house mouse) ] Gene ID: 66594, updated on 12-Aug-2019 Gene summary Official Symbol Uqcr11 provided by MGI Official Full Name ubiquinol-cytochrome c reductase, complex III subunit XI provided by MGI Primary source MGI:MGI:1913844 See related Ensembl:ENSMUSG00000020163 Gene type protein coding RefSeq status VALIDATED Organism Mus musculus Lineage Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi; Mammalia; Eutheria; Euarchontoglires; Glires; Rodentia; Myomorpha; Muroidea; Muridae; Murinae; Mus; Mus Also known as Uqcr; AL022707; 0710008D09Rik Expression Ubiquitous expression in heart adult (RPKM 1055.3), kidney adult (RPKM 899.6) and 28 other tissues See more Orthologs human all Genomic context Location: 10; 10 C1 See Uqcr11 in Genome Data Viewer Exon count: 3 Annotation release Status Assembly Chr Location 108 current GRCm38.p6 (GCF_000001635.26) 10 NC_000076.6 (80402997..80406821, complement) Build 37.2 previous assembly MGSCv37 (GCF_000001635.18) 10 NC_000076.5 (79865742..79869566, complement) Chromosome 10 - NC_000076.6 Page 5 of 8 https://www.alphaknockout.com Transcript information: This gene has 3 transcripts Gene: Uqcr11 ENSMUSG00000020163 Description ubiquinol-cytochrome c reductase, complex III subunit XI [Source:MGI Symbol;Acc:MGI:1913844] Gene Synonyms 0710008D09Rik, Uqcr Location Chromosome 10: 80,402,997-80,406,830 reverse strand. GRCm38:CM001003.2 About this gene This gene has 3 transcripts (splice variants), 150 orthologues and is a member of 1 Ensembl protein family. Transcripts Name Transcript ID bp Protein Translation ID Biotype CCDS UniProt Flags Uqcr11-201 ENSMUST00000020372.5 445 56aa ENSMUSP00000020372.5 Protein coding CCDS35981 Q9CPX8 TSL:1 GENCODE basic APPRIS P1 Uqcr11-203 ENSMUST00000141683.1 605 No protein - lncRNA - - TSL:1 Uqcr11-202 ENSMUST00000138428.7 520 No protein - lncRNA - - TSL:3 Page 6 of 8 https://www.alphaknockout.com 23.83 kb Forward strand 80.395Mb 80.400Mb 80.405Mb 80.410Mb 80.415Mb Contigs < AC152062.6 Genes (Comprehensive set... < Mbd3-204protein coding < Uqcr11-201protein coding < Tcf3-209protein coding < Mbd3-201protein coding < Uqcr11-202lncRNA < Tcf3-208protein coding < Mbd3-206protein coding < Uqcr11-203lncRNA < Tcf3-207protein coding < Mbd3-203protein coding < Tcf3-206protein coding < Mbd3-202protein coding < Tcf3-201protein coding < Mbd3-205lncRNA < Tcf3-210protein coding < Tcf3-205protein coding < Tcf3-202protein coding < Tcf3-204protein coding < Tcf3-203protein coding < Tcf3-214protein coding < Tcf3-212lncRNA < Tcf3-211retained intron Regulatory Build 80.395Mb 80.400Mb 80.405Mb 80.410Mb 80.415Mb Reverse strand 23.83 kb Regulation Legend CTCF Open Chromatin Promoter Promoter Flank Gene Legend Protein Coding merged Ensembl/Havana Ensembl protein coding Non-Protein Coding processed transcript RNA gene Page 7 of 8 https://www.alphaknockout.com Transcript: ENSMUST00000020372 < Uqcr11-201protein coding Reverse strand 3.83 kb ENSMUSP00000020... Transmembrane heli... Superfamily Single alpha-helix domain superfamily Pfam Cytochrome b-c1 complex subunit 10 PANTHER Cytochrome b-c1 complex subunit 10 Gene3D 1.20.5.220 Scale bar 0 6 12 18 24 30 36 42 48 56 We wish to acknowledge the following valuable scientific information resources: Ensembl, MGI, NCBI, UCSC. Page 8 of 8.
Load more

Recommended publications
  • Gene Expression Profiling Analysis of Lung Adenocarcinoma
    Brazilian Journal of Medical and Biological Research (2016) 49(3): e4861, http://dx.doi.org/10.1590/1414-431X20154861 ISSN 1414-431X 1/11 Gene expression profiling analysis of lung adenocarcinoma H. Xu1,2,J.Ma1,J.Wu1, L. Chen1,F.Sun1,C.Qu1, D. Zheng1 and S. Xu1 1Department of Thoracic Surgery, Harbin Medical University Cancer Hospital, Harbin, Heilongjiang, China 2Laboratory of Medical Genetics, Harbin Medical University, Harbin, Heilongjiang, China Abstract The present study screened potential genes related to lung adenocarcinoma, with the aim of further understanding disease pathogenesis. The GSE2514 dataset including 20 lung adenocarcinoma and 19 adjacent normal tissue samples from 10 patients with lung adenocarcinoma aged 45-73 years was downloaded from Gene Expression Omnibus. Differentially expressed genes (DEGs) between the two groups were screened using the t-test. Potential gene functions were predicted using functional and pathway enrichment analysis, and protein-protein interaction (PPI) networks obtained from the STRING database were constructed with Cytoscape. Module analysis of PPI networks was performed through MCODE in Cytoscape. In total, 535 upregulated and 465 downregulated DEGs were identified. These included ATP5D, UQCRC2, UQCR11 and genes encoding nicotinamide adenine dinucleotide (NADH), which are mainly associated with mitochondrial ATP synthesis coupled electron transport, and which were enriched in the oxidative phosphorylation pathway. Other DEGs were associated with DNA replication (PRIM1, MCM3, and RNASEH2A), cell surface receptor-linked signal transduction and the enzyme-linked receptor protein signaling pathway (MAPK1, STAT3, RAF1, and JAK1), and regulation of the cytoskeleton and phosphatidylinositol signaling system (PIP5K1B, PIP5K1C, and PIP4K2B). Our findings suggest that DEGs encoding subunits of NADH, PRIM1, MCM3, MAPK1, STAT3, RAF1, and JAK1 might be associated with the development of lung adenocarcinoma.
    [Show full text]
  • A High-Throughput Approach to Uncover Novel Roles of APOBEC2, a Functional Orphan of the AID/APOBEC Family
    Rockefeller University Digital Commons @ RU Student Theses and Dissertations 2018 A High-Throughput Approach to Uncover Novel Roles of APOBEC2, a Functional Orphan of the AID/APOBEC Family Linda Molla Follow this and additional works at: https://digitalcommons.rockefeller.edu/ student_theses_and_dissertations Part of the Life Sciences Commons A HIGH-THROUGHPUT APPROACH TO UNCOVER NOVEL ROLES OF APOBEC2, A FUNCTIONAL ORPHAN OF THE AID/APOBEC FAMILY A Thesis Presented to the Faculty of The Rockefeller University in Partial Fulfillment of the Requirements for the degree of Doctor of Philosophy by Linda Molla June 2018 © Copyright by Linda Molla 2018 A HIGH-THROUGHPUT APPROACH TO UNCOVER NOVEL ROLES OF APOBEC2, A FUNCTIONAL ORPHAN OF THE AID/APOBEC FAMILY Linda Molla, Ph.D. The Rockefeller University 2018 APOBEC2 is a member of the AID/APOBEC cytidine deaminase family of proteins. Unlike most of AID/APOBEC, however, APOBEC2’s function remains elusive. Previous research has implicated APOBEC2 in diverse organisms and cellular processes such as muscle biology (in Mus musculus), regeneration (in Danio rerio), and development (in Xenopus laevis). APOBEC2 has also been implicated in cancer. However the enzymatic activity, substrate or physiological target(s) of APOBEC2 are unknown. For this thesis, I have combined Next Generation Sequencing (NGS) techniques with state-of-the-art molecular biology to determine the physiological targets of APOBEC2. Using a cell culture muscle differentiation system, and RNA sequencing (RNA-Seq) by polyA capture, I demonstrated that unlike the AID/APOBEC family member APOBEC1, APOBEC2 is not an RNA editor. Using the same system combined with enhanced Reduced Representation Bisulfite Sequencing (eRRBS) analyses I showed that, unlike the AID/APOBEC family member AID, APOBEC2 does not act as a 5-methyl-C deaminase.
    [Show full text]
  • Lncrna SNHG8 Is Identified As a Key Regulator of Acute Myocardial
    Zhuo et al. Lipids in Health and Disease (2019) 18:201 https://doi.org/10.1186/s12944-019-1142-0 RESEARCH Open Access LncRNA SNHG8 is identified as a key regulator of acute myocardial infarction by RNA-seq analysis Liu-An Zhuo, Yi-Tao Wen, Yong Wang, Zhi-Fang Liang, Gang Wu, Mei-Dan Nong and Liu Miao* Abstract Background: Long noncoding RNAs (lncRNAs) are involved in numerous physiological functions. However, their mechanisms in acute myocardial infarction (AMI) are not well understood. Methods: We performed an RNA-seq analysis to explore the molecular mechanism of AMI by constructing a lncRNA-miRNA-mRNA axis based on the ceRNA hypothesis. The target microRNA data were used to design a global AMI triple network. Thereafter, a functional enrichment analysis and clustering topological analyses were conducted by using the triple network. The expression of lncRNA SNHG8, SOCS3 and ICAM1 was measured by qRT-PCR. The prognostic values of lncRNA SNHG8, SOCS3 and ICAM1 were evaluated using a receiver operating characteristic (ROC) curve. Results: An AMI lncRNA-miRNA-mRNA network was constructed that included two mRNAs, one miRNA and one lncRNA. After RT-PCR validation of lncRNA SNHG8, SOCS3 and ICAM1 between the AMI and normal samples, only lncRNA SNHG8 had significant diagnostic value for further analysis. The ROC curve showed that SNHG8 presented an AUC of 0.850, while the AUC of SOCS3 was 0.633 and that of ICAM1 was 0.594. After a pairwise comparison, we found that SNHG8 was statistically significant (P SNHG8-ICAM1 = 0.002; P SNHG8-SOCS3 = 0.031).
    [Show full text]
  • Human Mitochondrial Pathologies of the Respiratory Chain and ATP Synthase: Contributions from Studies of Saccharomyces Cerevisiae
    life Review Human Mitochondrial Pathologies of the Respiratory Chain and ATP Synthase: Contributions from Studies of Saccharomyces cerevisiae Leticia V. R. Franco 1,2,* , Luca Bremner 1 and Mario H. Barros 2 1 Department of Biological Sciences, Columbia University, New York, NY 10027, USA; [email protected] 2 Department of Microbiology,Institute of Biomedical Sciences, Universidade de Sao Paulo, Sao Paulo 05508-900, Brazil; [email protected] * Correspondence: [email protected] Received: 27 October 2020; Accepted: 19 November 2020; Published: 23 November 2020 Abstract: The ease with which the unicellular yeast Saccharomyces cerevisiae can be manipulated genetically and biochemically has established this organism as a good model for the study of human mitochondrial diseases. The combined use of biochemical and molecular genetic tools has been instrumental in elucidating the functions of numerous yeast nuclear gene products with human homologs that affect a large number of metabolic and biological processes, including those housed in mitochondria. These include structural and catalytic subunits of enzymes and protein factors that impinge on the biogenesis of the respiratory chain. This article will review what is currently known about the genetics and clinical phenotypes of mitochondrial diseases of the respiratory chain and ATP synthase, with special emphasis on the contribution of information gained from pet mutants with mutations in nuclear genes that impair mitochondrial respiration. Our intent is to provide the yeast mitochondrial specialist with basic knowledge of human mitochondrial pathologies and the human specialist with information on how genes that directly and indirectly affect respiration were identified and characterized in yeast. Keywords: mitochondrial diseases; respiratory chain; yeast; Saccharomyces cerevisiae; pet mutants 1.
    [Show full text]
  • Electron Transport Chain Activity Is a Predictor and Target for Venetoclax Sensitivity in Multiple Myeloma
    ARTICLE https://doi.org/10.1038/s41467-020-15051-z OPEN Electron transport chain activity is a predictor and target for venetoclax sensitivity in multiple myeloma Richa Bajpai1,7, Aditi Sharma 1,7, Abhinav Achreja2,3, Claudia L. Edgar1, Changyong Wei1, Arusha A. Siddiqa1, Vikas A. Gupta1, Shannon M. Matulis1, Samuel K. McBrayer 4, Anjali Mittal3,5, Manali Rupji 6, Benjamin G. Barwick 1, Sagar Lonial1, Ajay K. Nooka 1, Lawrence H. Boise 1, Deepak Nagrath2,3,5 & ✉ Mala Shanmugam 1 1234567890():,; The BCL-2 antagonist venetoclax is highly effective in multiple myeloma (MM) patients exhibiting the 11;14 translocation, the mechanistic basis of which is unknown. In evaluating cellular energetics and metabolism of t(11;14) and non-t(11;14) MM, we determine that venetoclax-sensitive myeloma has reduced mitochondrial respiration. Consistent with this, low electron transport chain (ETC) Complex I and Complex II activities correlate with venetoclax sensitivity. Inhibition of Complex I, using IACS-010759, an orally bioavailable Complex I inhibitor in clinical trials, as well as succinate ubiquinone reductase (SQR) activity of Complex II, using thenoyltrifluoroacetone (TTFA) or introduction of SDHC R72C mutant, independently sensitize resistant MM to venetoclax. We demonstrate that ETC inhibition increases BCL-2 dependence and the ‘primed’ state via the ATF4-BIM/NOXA axis. Further, SQR activity correlates with venetoclax sensitivity in patient samples irrespective of t(11;14) status. Use of SQR activity in a functional-biomarker informed manner may better select for MM patients responsive to venetoclax therapy. 1 Department of Hematology and Medical Oncology, Winship Cancer Institute, School of Medicine, Emory University, Atlanta, GA, USA.
    [Show full text]
  • Primepcr™Assay Validation Report
    PrimePCR™Assay Validation Report Gene Information Gene Name ubiquinol-cytochrome c reductase, complex III subunit XI Gene Symbol Uqcr11 Organism Mouse Gene Summary Description Not Available Gene Aliases 0710008D09Rik, AL022707, Uqcr RefSeq Accession No. NC_000076.6, NT_039500.8 UniGene ID Mm.379119 Ensembl Gene ID ENSMUSG00000020163 Entrez Gene ID 66594 Assay Information Unique Assay ID qMmuCEP0053751 Assay Type Probe - Validation information is for the primer pair using SYBR® Green detection Detected Coding Transcript(s) ENSMUST00000020372 Amplicon Context Sequence AGTGTCTGTCAACATGGAAGCTCCCTGAGCGGTGCAGACCTCTCCAGTGTCCAG CTTCCTCAGTGTCTTCCACAGGACTACGCTGTGTCTGAGGCCTCGGCACATGG Amplicon Length (bp) 77 Chromosome Location 10:80403046-80403152 Assay Design Exonic Purification Desalted Validation Results Efficiency (%) 97 R2 0.9999 cDNA Cq 18.22 cDNA Tm (Celsius) 83 gDNA Cq 25.26 Specificity (%) 100 Information to assist with data interpretation is provided at the end of this report. Page 1/4 PrimePCR™Assay Validation Report Uqcr11, Mouse Amplification Plot Amplification of cDNA generated from 25 ng of universal reference RNA Melt Peak Melt curve analysis of above amplification Standard Curve Standard curve generated using 20 million copies of template diluted 10-fold to 20 copies Page 2/4 PrimePCR™Assay Validation Report Products used to generate validation data Real-Time PCR Instrument CFX384 Real-Time PCR Detection System Reverse Transcription Reagent iScript™ Advanced cDNA Synthesis Kit for RT-qPCR Real-Time PCR Supermix SsoAdvanced™ SYBR® Green Supermix Experimental Sample qPCR Mouse Reference Total RNA Data Interpretation Unique Assay ID This is a unique identifier that can be used to identify the assay in the literature and online. Detected Coding Transcript(s) This is a list of the Ensembl transcript ID(s) that this assay will detect.
    [Show full text]
  • Downloaded Per Proteome Cohort Via the Web- Site Links of Table 1, Also Providing Information on the Deposited Spectral Datasets
    www.nature.com/scientificreports OPEN Assessment of a complete and classifed platelet proteome from genome‑wide transcripts of human platelets and megakaryocytes covering platelet functions Jingnan Huang1,2*, Frauke Swieringa1,2,9, Fiorella A. Solari2,9, Isabella Provenzale1, Luigi Grassi3, Ilaria De Simone1, Constance C. F. M. J. Baaten1,4, Rachel Cavill5, Albert Sickmann2,6,7,9, Mattia Frontini3,8,9 & Johan W. M. Heemskerk1,9* Novel platelet and megakaryocyte transcriptome analysis allows prediction of the full or theoretical proteome of a representative human platelet. Here, we integrated the established platelet proteomes from six cohorts of healthy subjects, encompassing 5.2 k proteins, with two novel genome‑wide transcriptomes (57.8 k mRNAs). For 14.8 k protein‑coding transcripts, we assigned the proteins to 21 UniProt‑based classes, based on their preferential intracellular localization and presumed function. This classifed transcriptome‑proteome profle of platelets revealed: (i) Absence of 37.2 k genome‑ wide transcripts. (ii) High quantitative similarity of platelet and megakaryocyte transcriptomes (R = 0.75) for 14.8 k protein‑coding genes, but not for 3.8 k RNA genes or 1.9 k pseudogenes (R = 0.43–0.54), suggesting redistribution of mRNAs upon platelet shedding from megakaryocytes. (iii) Copy numbers of 3.5 k proteins that were restricted in size by the corresponding transcript levels (iv) Near complete coverage of identifed proteins in the relevant transcriptome (log2fpkm > 0.20) except for plasma‑derived secretory proteins, pointing to adhesion and uptake of such proteins. (v) Underrepresentation in the identifed proteome of nuclear‑related, membrane and signaling proteins, as well proteins with low‑level transcripts.
    [Show full text]
  • Molecular Architecture Underlying Fluid Absorption by the Developing Inner
    RESEARCH ARTICLE Molecular architecture underlying fluid absorption by the developing inner ear Keiji Honda1†, Sung Huhn Kim2‡, Michael C Kelly3, Joseph C Burns3§, Laura Constance2, Xiangming Li2#, Fei Zhou2, Michael Hoa4, Matthew W Kelley3, Philine Wangemann2*, Robert J Morell5, Andrew J Griffith1* 1Molecular Biology and Genetics Section, National Institute on Deafness and Other Communication Disorders, National Institutes of Health, Bethesda, United States; 2Anatomy and Physiology Department, Kansas State University, Manhattan, United States; 3Developmental Neuroscience Section, National Institute on Deafness and Other Communication Disorders, National Institutes of Health, Bethesda, United States; 4Auditory Development and Restoration Program, National Institute on *For correspondence: Deafness and Other Communication Disorders, National Institutes of Health, [email protected] (PW); Bethesda, United States; 5Genomics and Computational Biology Core, National [email protected] (AJG) Institute on Deafness and Other Communication Disorders, National Institutes of Present address: Health, Bethesda, United States †Otolaryngology Department, Tsuchiura Kyodo General Hospital, Tsuchiura, Japan; ‡Department of Abstract Mutations of SLC26A4 are a common cause of hearing loss associated with Otorhinolaryngology, Head and enlargement of the endolymphatic sac (EES). Slc26a4 expression in the developing mouse Neck Surgery, Yonsei University College of Medicine, Seoul, endolymphatic sac is required for acquisition of normal inner ear structure and function. Here, we Korea; §Decibel Therapeutics, show that the mouse endolymphatic sac absorbs fluid in an SLC26A4-dependent fashion. Fluid Cambridge, United States; absorption was sensitive to ouabain and gadolinium but insensitive to benzamil, bafilomycin and #Technique R and D-Drug S3226. Single-cell RNA-seq analysis of pre- and postnatal endolymphatic sacs demonstrates two Substance, GlaxoSmithKline types of differentiated cells.
    [Show full text]
  • Mitochondrial Structure and Bioenergetics in Normal and Disease Conditions
    International Journal of Molecular Sciences Review Mitochondrial Structure and Bioenergetics in Normal and Disease Conditions Margherita Protasoni 1 and Massimo Zeviani 1,2,* 1 Mitochondrial Biology Unit, The MRC and University of Cambridge, Cambridge CB2 0XY, UK; [email protected] 2 Department of Neurosciences, University of Padova, 35128 Padova, Italy * Correspondence: [email protected] Abstract: Mitochondria are ubiquitous intracellular organelles found in almost all eukaryotes and involved in various aspects of cellular life, with a primary role in energy production. The interest in this organelle has grown stronger with the discovery of their link to various pathologies, including cancer, aging and neurodegenerative diseases. Indeed, dysfunctional mitochondria cannot provide the required energy to tissues with a high-energy demand, such as heart, brain and muscles, leading to a large spectrum of clinical phenotypes. Mitochondrial defects are at the origin of a group of clinically heterogeneous pathologies, called mitochondrial diseases, with an incidence of 1 in 5000 live births. Primary mitochondrial diseases are associated with genetic mutations both in nuclear and mitochondrial DNA (mtDNA), affecting genes involved in every aspect of the organelle function. As a consequence, it is difficult to find a common cause for mitochondrial diseases and, subsequently, to offer a precise clinical definition of the pathology. Moreover, the complexity of this condition makes it challenging to identify possible therapies or drug targets. Keywords: ATP production; biogenesis of the respiratory chain; mitochondrial disease; mi-tochondrial electrochemical gradient; mitochondrial potential; mitochondrial proton pumping; mitochondrial respiratory chain; oxidative phosphorylation; respiratory complex; respiratory supercomplex Citation: Protasoni, M.; Zeviani, M.
    [Show full text]
  • UQCR11 (NM 006830) Human Tagged ORF Clone – RC201360
    OriGene Technologies, Inc. 9620 Medical Center Drive, Ste 200 Rockville, MD 20850, US Phone: +1-888-267-4436 [email protected] EU: [email protected] CN: [email protected] Product datasheet for RC201360 UQCR11 (NM_006830) Human Tagged ORF Clone Product data: Product Type: Expression Plasmids Product Name: UQCR11 (NM_006830) Human Tagged ORF Clone Tag: Myc-DDK Symbol: UQCR11 Synonyms: 0710008D09Rik; QCR10; UQCR Vector: pCMV6-Entry (PS100001) E. coli Selection: Kanamycin (25 ug/mL) Cell Selection: Neomycin ORF Nucleotide >RC201360 representing NM_006830 Sequence: Red=Cloning site Blue=ORF Green=Tags(s) TTTTGTAATACGACTCACTATAGGGCGGCCGGGAATTCGTCGACTGGATCCGGTACCGAGGAGATCTGCC GCCGCGATCGCC ATGGTGACCCGGTTCCTGGGCCCACGCTACCGGGAGCTGGTCAAGAACTGGGTCCCGACGGCCTACACAT GGGGCGCTGTGGGCGCCGTGGGGCTGGTGTGGGCCACCGATTGGCGGCTGATCCTGGACTGGGTACCTTA CATCAATGGCAAGTTTAAGAAGGATAAT ACGCGTACGCGGCCGCTCGAGCAGAAACTCATCTCAGAAGAGGATCTGGCAGCAAATGATATCCTGGATT ACAAGGATGACGACGATAAGGTTTAA Protein Sequence: >RC201360 representing NM_006830 Red=Cloning site Green=Tags(s) MVTRFLGPRYRELVKNWVPTAYTWGAVGAVGLVWATDWRLILDWVPYINGKFKKDN TRTRPLEQKLISEEDLAANDILDYKDDDDKV Chromatograms: https://cdn.origene.com/chromatograms/mk8112_c05.zip Restriction Sites: SgfI-MluI This product is to be used for laboratory only. Not for diagnostic or therapeutic use. View online » ©2021 OriGene Technologies, Inc., 9620 Medical Center Drive, Ste 200, Rockville, MD 20850, US 1 / 3 UQCR11 (NM_006830) Human Tagged ORF Clone – RC201360 Cloning Scheme: Plasmid Map: ACCN: NM_006830 ORF Size: 168 bp This
    [Show full text]
  • UQCRC2 Polyclonal Antibody
    For Research Use Only UQCRC2 Polyclonal antibody Catalog Number:14742-1-AP Featured Product 26 Publications www.ptgcn.com Catalog Number: GenBank Accession Number: Recommended Dilutions: Basic Information 14742-1-AP BC000484 WB 1:2000-1:12000 Size: GeneID (NCBI): IP 0.5-4.0 ug for IP and 1:500-1:2000 327 μg/ml 7385 for WB IHC 1:50-1:500 Source: Full Name: IF 1:50-1:500 Rabbit ubiquinol-cytochrome c reductase Isotype: core protein II IgG Calculated MW: Purification Method: 48 kDa Antigen affinity purification Observed MW: Immunogen Catalog Number: 48 kDa AG6432 Applications Tested Applications: Positive Controls: FC, IF, IHC, IP, WB, ELISA WB : mouse brain tissue; human colon tissue, mouse Cited Applications: colon tissue, mouse heart tissue, mouse placenta IF, IHC, WB tissue, mouse skeletal muscle tissue, SGC-7901 cells, Species Specificity: various samples, rat heart tissue human, mouse, rat IP : mouse brain tissue; Cited Species: IHC : human colon cancer tissue; human, mouse, rat IF : HepG2 cells; Note-IHC: suggested antigen retrieval with TE buffer pH 9.0; (*) Alternatively, antigen retrieval may be performed with citrate buffer pH 6.0 UQCRC2(Ubiquinol-cytochrome-c reductase complex core protein 2) is also named as complex III subunit 2,core Background Information protein II and belongs to the UQCRC2/QCR2 subfamily. The bc1 complex contains 11 subunits: 3 respiratory subunits (cytochrome b, cytochrome c1 and Rieske/UQCRFS1), 2 core proteins (UQCRC1/QCR1 and UQCRC2/QCR2) and 6 low-molecular weight proteins (UQCRH/QCR6, UQCRB/QCR7, UQCRQ/QCR8, UQCR10/QCR9, UQCR11/QCR10 and a cleavage product of Rieske/UQCRFS1) and is part of the mitochondrial respiratory chain.
    [Show full text]
  • Nuclear Gene Mutations As the Cause of Mitochondrial Complex III Deficiency
    REVIEW published: 09 April 2015 doi: 10.3389/fgene.2015.00134 Nuclear gene mutations as the cause of mitochondrial complex III deficiency Erika Fernández-Vizarra*† and Massimo Zeviani Mitochondrial Biology Unit, Medical Research Council, Cambridge, UK Complex III (CIII) deficiency is one of the least common oxidative phosphorylation defects associated to mitochondrial disease. CIII constitutes the center of the mitochondrial respiratory chain, as well as a crossroad for several other metabolic pathways. For more than 10 years, of all the potential candidate genes encoding structural subunits and assembly factors, only three were known to be associated to CIII defects in human pathology. Thus, leaving many of these cases unresolved. These first identified Edited by: genes were MT-CYB, the only CIII subunit encoded in the mitochondrial DNA; BCS1L, Tiziana Lodi, encoding an assembly factor, and UQCRB, a nuclear-encoded structural subunit. University of Parma, Italy Reviewed by: Nowadays, thanks to the fast progress that has taken place in the last 3–4 years, Saima Siddiqi, pathological changes in seven more genes are known to be associated to these Institute of Biomedical and Genetic conditions. This review will focus on the strategies that have permitted the latest Engineering, Pakistan Vineta Fellman, discovery of mutations in factors that are necessary for a correct CIII assembly and Lund University, Sweden activity, in relation with their function. In addition, new data further establishing the *Correspondence: molecular role of LYRM7/MZM1L
    [Show full text]