https://www.alphaknockout.com
Mouse Rnf135 Knockout Project (CRISPR/Cas9)
Objective: To create a Rnf135 knockout Mouse model (C57BL/6J) by CRISPR/Cas-mediated genome engineering.
Strategy summary: The Rnf135 gene (NCBI Reference Sequence: NM_028019 ; Ensembl: ENSMUSG00000020707 ) is located on Mouse chromosome 11. 5 exons are identified, with the ATG start codon in exon 1 and the TGA stop codon in exon 5 (Transcript: ENSMUST00000017839). Exon 2~4 will be selected as target site. Cas9 and gRNA will be co-injected into fertilized eggs for KO Mouse production. The pups will be genotyped by PCR followed by sequencing analysis. Note:
Exon 2 starts from about 27.18% of the coding region. Exon 2~4 covers 30.54% of the coding region. The size of effective KO region: ~7724 bp. The KO region does not have any other known gene.
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Overview of the Targeting Strategy
Wildtype allele 5' gRNA region gRNA region 3'
1 2 3 4 5
Legends Exon of mouse Rnf135 Knockout region
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Overview of the Dot Plot (up) Window size: 15 bp
Forward Reverse Complement
Sequence 12
Note: The 2000 bp section upstream of Exon 2 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 Dot Plot (down) Window size: 15 bp
Forward Reverse Complement
Sequence 12
Note: The 1613 bp section downstream of Exon 4 is aligned with itself to determine if there are tandem repeats. Tandem repeats are found in the dot plot matrix. The gRNA site is selected outside of these tandem repeats.
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Overview of the GC Content Distribution (up) Window size: 300 bp
Sequence 12
Summary: Full Length(2000bp) | A(27.35% 547) | C(22.45% 449) | T(31.0% 620) | G(19.2% 384)
Note: The 2000 bp section upstream of Exon 2 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.
Overview of the GC Content Distribution (down) Window size: 300 bp
Sequence 12
Summary: Full Length(1613bp) | A(25.91% 418) | C(22.88% 369) | T(29.94% 483) | G(21.26% 343)
Note: The 1613 bp section downstream of Exon 4 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.
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BLAT Search Results (up)
QUERY SCORE START END QSIZE IDENTITY CHROM STRAND START END SPAN ------browser details YourSeq 2000 1 2000 2000 100.0% chr11 + 80187225 80189224 2000 browser details YourSeq 1053 34 1325 2000 96.8% chr8 - 99657964 99659215 1252 browser details YourSeq 1031 24 1089 2000 98.5% chr4 - 113968946 114237216 268271 browser details YourSeq 1030 23 1089 2000 98.7% chr11 - 8742765 8743835 1071 browser details YourSeq 1029 25 1089 2000 98.9% chr11 + 10223478 10224549 1072 browser details YourSeq 1028 24 1089 2000 98.0% chr18 - 54417560 54418614 1055 browser details YourSeq 1028 25 1089 2000 98.7% chr1 - 9715368 9716437 1070 browser details YourSeq 1027 26 1089 2000 98.1% chr1 - 79689410 79690462 1053 browser details YourSeq 1027 31 1089 2000 98.4% chr9 + 89321374 89322416 1043 browser details YourSeq 1026 23 1089 2000 98.5% chr12 - 63109565 63110634 1070 browser details YourSeq 1025 23 1089 2000 97.9% chr10 - 12050103 12051155 1053 browser details YourSeq 1025 35 1089 2000 98.3% chr1 - 154001149 154002194 1046 browser details YourSeq 1025 29 1089 2000 98.1% chr5 + 73587593 73588643 1051 browser details YourSeq 1025 33 1089 2000 98.2% chr16 + 12621855 12622901 1047 browser details YourSeq 1025 28 1089 2000 98.2% chr13 + 78607251 78608308 1058 browser details YourSeq 1025 36 1089 2000 98.4% chr13 + 30273538 30274581 1044 browser details YourSeq 1024 24 1089 2000 98.2% chr7 - 144727049 144728109 1061 browser details YourSeq 1024 26 1089 2000 97.9% chr10 - 34375086 34376137 1052 browser details YourSeq 1024 34 1089 2000 98.2% chr5 + 107575602 107576648 1047 browser details YourSeq 1023 25 1089 2000 98.2% chr3 - 158574112 158575174 1063
Note: The 2000 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 1613 1 1613 1613 100.0% chr11 + 80196949 80198561 1613 browser details YourSeq 127 351 1433 1613 92.1% chr14 - 70308280 70704212 395933 browser details YourSeq 119 351 1432 1613 95.5% chr1 + 131927564 132101080 173517 browser details YourSeq 117 353 1430 1613 94.0% chr8 + 84938407 84948048 9642 browser details YourSeq 106 351 1432 1613 92.8% chr17 + 15668576 15869416 200841 browser details YourSeq 92 351 1407 1613 94.3% chr11 + 62376285 62589001 212717 browser details YourSeq 90 351 455 1613 93.4% chr11 - 69891985 69892254 270 browser details YourSeq 87 144 420 1613 86.6% chr10 + 81220000 81220454 455 browser details YourSeq 86 351 451 1613 94.8% chr2 - 8455031 8455147 117 browser details YourSeq 84 351 455 1613 97.8% chrX - 89480391 89480511 121 browser details YourSeq 77 329 427 1613 93.3% chr12 - 54898164 54898393 230 browser details YourSeq 74 346 427 1613 95.2% chr10 + 26069950 26070031 82 browser details YourSeq 73 351 427 1613 97.5% chr5 - 87929121 87929197 77 browser details YourSeq 73 351 427 1613 97.5% chr19 - 5399372 5399448 77 browser details YourSeq 73 351 427 1613 97.5% chr12 - 60808991 60809067 77 browser details YourSeq 73 351 427 1613 97.5% chr11 - 96094576 96094652 77 browser details YourSeq 73 351 427 1613 97.5% chr8 + 93931050 93931126 77 browser details YourSeq 73 351 427 1613 97.5% chr18 + 57268215 57268291 77 browser details YourSeq 73 351 427 1613 97.5% chr17 + 29289140 29289216 77 browser details YourSeq 73 351 427 1613 97.5% chr12 + 83966661 83966737 77
Note: The 1613 bp section downstream of Exon 4 is BLAT searched against the genome. No significant similarity is found.
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Gene and protein information: Rnf135 ring finger protein 135 [ Mus musculus (house mouse) ] Gene ID: 71956, updated on 12-Aug-2019
Gene summary
Official Symbol Rnf135 provided by MGI Official Full Name ring finger protein 135 provided by MGI Primary source MGI:MGI:1919206 See related Ensembl:ENSMUSG00000020707 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 Riplet; U 2-3-0; 0610037N03Rik; 2410006N06Rik Expression Ubiquitous expression in ovary adult (RPKM 5.9), adrenal adult (RPKM 5.2) and 28 other tissues See more Orthologs human all
Genomic context
Location: 11 B5; 11 47.59 cM See Rnf135 in Genome Data Viewer Exon count: 5
Annotation release Status Assembly Chr Location
108 current GRCm38.p6 (GCF_000001635.26) 11 NC_000077.6 (80183869..80199755)
Build 37.2 previous assembly MGSCv37 (GCF_000001635.18) 11 NC_000077.5 (79997374..80013255)
Chromosome 11 - NC_000077.6
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Transcript information: This gene has 2 transcripts
Gene: Rnf135 ENSMUSG00000020707
Description ring finger protein 135 [Source:MGI Symbol;Acc:MGI:1919206] Gene Synonyms 0610037N03Rik, 2410006N06Rik, MGC13061, U 2-3-0 Location Chromosome 11: 80,183,851-80,199,757 forward strand. GRCm38:CM001004.2 About this gene This gene has 2 transcripts (splice variants), 84 orthologues, 73 paralogues, is a member of 1 Ensembl protein family and is associated with 4 phenotypes. Transcripts
Name Transcript ID bp Protein Translation ID Biotype CCDS UniProt Flags
Rnf135- ENSMUST00000017839.2 1984 417aa ENSMUSP00000017839.2 Protein CCDS25129 B2RRA5 Q9CWS1 TSL:1 201 coding GENCODE basic APPRIS P1
Rnf135- ENSMUST00000134909.1 333 No - lncRNA - - TSL:5 202 protein
35.91 kb Forward strand
80.18Mb 80.19Mb 80.20Mb Genes (Comprehensive set... Adap2-201 >protein coding Rnf135-201 >protein coding Rhot1-209 >retained intron
Rnf135-202 >lncRNA Rhot1-205 >retained intron
Rhot1-203 >protein coding
Rhot1-202 >protein coding
Rhot1-204 >protein coding
Rhot1-201 >protein coding
Contigs AL663057.7 > AL591426.17 > Genes < Adap2os-201lncRNA (Comprehensive set...
Regulatory Build
80.18Mb 80.19Mb 80.20Mb Reverse strand 35.91 kb
Regulation Legend CTCF Open Chromatin Promoter Promoter Flank
Gene Legend Protein Coding
Ensembl protein coding merged Ensembl/Havana
Non-Protein Coding
processed transcript RNA gene
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Transcript: ENSMUST00000017839
15.91 kb Forward strand
Rnf135-201 >protein coding
ENSMUSP00000017... MobiDB lite Low complexity (Seg) Coiled-coils (Ncoils) Superfamily SSF57850 Concanavalin A-like lectin/glucanase domain superfamily
SMART Zinc finger, RING-type SPRY domain
Prints Butyrophylin-like, SPRY domain Pfam PF15227 SPRY domain
PROSITE profiles Zinc finger, RING-type B30.2/SPRY domain
PROSITE patterns Zinc finger, RING-type, conserved site PANTHER PTHR25465:SF16
PTHR25465 Gene3D Zinc finger, RING/FYVE/PHD-type 2.60.120.920
CDD cd12902
All sequence SNPs/i... Sequence variants (dbSNP and all other sources)
Variant Legend frameshift variant missense variant synonymous variant
Scale bar 0 40 80 120 160 200 240 280 320 360 417
We wish to acknowledge the following valuable scientific information resources: Ensembl, MGI, NCBI, UCSC.
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