https://www.alphaknockout.com
Mouse Trim27 Conditional Knockout Project (CRISPR/Cas9)
Objective: To create a Trim27 conditional knockout Mouse model (C57BL/6J) by CRISPR/Cas-mediated genome engineering.
Strategy summary: The Trim27 gene (NCBI Reference Sequence: NM_009054 ; Ensembl: ENSMUSG00000021326 ) is located on Mouse chromosome 13. 8 exons are identified, with the ATG start codon in exon 1 and the TGA stop codon in exon 8 (Transcript: ENSMUST00000021761). Exon 4~6 will be selected as conditional knockout region (cKO region). Deletion of this region should result in the loss of function of the Mouse Trim27 gene. To engineer the targeting vector, homologous arms and cKO region will be generated by PCR using BAC clone RP23-421D22 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: Mice homozygous for a gene trap allele exhibit exhibit increased potassium/calcium channel activity and TCR- stimulated calcium influx in Th1 and Th2 CD4 T cells. Mice homozygous for another gene trap allele exhibit decreased incidence of chemically-induced tumors.
Exon 4 starts from about 48.6% of the coding region. The knockout of Exon 4~6 will result in frameshift of the gene. The size of intron 3 for 5'-loxP site insertion: 4078 bp, and the size of intron 6 for 3'-loxP site insertion: 1095 bp. The size of effective cKO region: ~2584 bp. The cKO 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 4 5 6 7 8 Targeting vector
Targeted allele
Constitutive KO allele (After Cre recombination)
Legends Exon of mouse Trim27 Homology arm cKO region loxP site
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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. Tandem repeats are found in the dot plot matrix. It may be difficult to construct this targeting vector.
Overview of the GC Content Distribution Window size: 300 bp
Sequence 12
Summary: Full Length(9084bp) | A(27.58% 2505) | C(19.31% 1754) | T(28.92% 2627) | G(24.2% 2198)
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.
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BLAT Search Results (up)
QUERY SCORE START END QSIZE IDENTITY CHROM STRAND START END SPAN ------browser details YourSeq 3000 1 3000 3000 100.0% chr13 + 21184792 21187791 3000 browser details YourSeq 136 1622 1776 3000 94.2% chr11 - 112087525 112087680 156 browser details YourSeq 133 1606 1777 3000 89.5% chrX + 155221046 155221220 175 browser details YourSeq 133 1607 1767 3000 92.9% chr10 + 122434369 122434529 161 browser details YourSeq 132 1622 1773 3000 92.1% chr2 + 108943067 108943217 151 browser details YourSeq 132 1622 1777 3000 89.6% chr10 + 124139018 124139170 153 browser details YourSeq 131 1607 1763 3000 90.4% chr1 + 153370696 153370851 156 browser details YourSeq 130 1626 1777 3000 90.7% chr11 + 31787198 31787347 150 browser details YourSeq 129 1631 1777 3000 96.5% chr10 - 26442964 26443113 150 browser details YourSeq 129 1622 1764 3000 93.7% chr19 + 7304597 7304738 142 browser details YourSeq 128 1605 1767 3000 91.6% chr14 - 8086902 8087065 164 browser details YourSeq 127 1622 1762 3000 93.6% chr11 - 78331314 78331453 140 browser details YourSeq 127 1622 1773 3000 93.8% chr10 - 45121437 45121588 152 browser details YourSeq 126 1622 1767 3000 91.8% chr4 - 98976580 98976724 145 browser details YourSeq 126 1622 1970 3000 92.0% chr15 - 99784332 99784762 431 browser details YourSeq 126 1621 1773 3000 91.4% chr8 + 96380391 96380543 153 browser details YourSeq 126 1626 1777 3000 89.6% chr5 + 43659244 43659390 147 browser details YourSeq 126 1621 1762 3000 93.0% chr11 + 11805259 11805399 141 browser details YourSeq 126 1622 1764 3000 94.4% chr1 + 76721510 76721652 143 browser details YourSeq 125 1632 1776 3000 93.8% chr8 - 69547136 69547289 154
Note: The 3000 bp section upstream of Exon 4 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% chr13 + 21190376 21193375 3000 browser details YourSeq 81 1652 2060 3000 65.4% chr13 - 23458965 23459370 406 browser details YourSeq 60 2608 2884 3000 90.3% chr11 + 21540291 21540647 357 browser details YourSeq 49 505 610 3000 91.6% chr10 - 18935559 18935673 115 browser details YourSeq 48 513 610 3000 92.9% chr17 - 46146723 46146822 100 browser details YourSeq 48 1366 1487 3000 87.5% chr10 + 39386715 39386834 120 browser details YourSeq 47 508 610 3000 91.3% chr5 - 92529035 92529137 103 browser details YourSeq 47 566 628 3000 91.3% chr18 - 66504614 66504677 64 browser details YourSeq 47 508 610 3000 91.3% chr13 - 90922126 90922228 103 browser details YourSeq 47 2872 2970 3000 88.6% chr14 + 29344556 29344656 101 browser details YourSeq 47 2872 2970 3000 88.6% chr1 + 119704229 119704329 101 browser details YourSeq 46 513 610 3000 94.3% chr2 - 119866305 119866402 98 browser details YourSeq 46 576 637 3000 85.0% chr11 - 68945999 68946059 61 browser details YourSeq 46 513 610 3000 91.1% chr17 + 64541496 64541593 98 browser details YourSeq 45 514 610 3000 94.2% chr1 + 44663575 44663671 97 browser details YourSeq 44 513 610 3000 92.4% chr7 - 90680311 90680408 98 browser details YourSeq 44 513 610 3000 92.4% chr11 + 102034110 102034207 98 browser details YourSeq 44 513 610 3000 92.4% chr11 + 70692510 70692607 98 browser details YourSeq 43 513 610 3000 92.2% chr10 - 97469116 97469213 98 browser details YourSeq 42 566 623 3000 90.4% chr11 - 6464011 6464071 61
Note: The 3000 bp section downstream of Exon 6 is BLAT searched against the genome. No significant similarity is found.
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Gene and protein information: Trim27 tripartite motif-containing 27 [ Mus musculus (house mouse) ] Gene ID: 19720, updated on 12-Aug-2019
Gene summary
Official Symbol Trim27 provided by MGI Official Full Name tripartite motif-containing 27 provided by MGI Primary source MGI:MGI:97904 See related Ensembl:ENSMUSG00000021326 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 Rfp; Gm19403; AW538890 Expression Ubiquitous expression in ovary adult (RPKM 33.3), adrenal adult (RPKM 32.0) and 28 other tissues See more Orthologs human all
Genomic context
Location: 13 A2; 13 7.67 cM See Trim27 in Genome Data Viewer
Exon count: 10
Annotation release Status Assembly Chr Location
108 current GRCm38.p6 (GCF_000001635.26) 13 NC_000079.6 (21179450..21194723)
Build 37.2 previous assembly MGSCv37 (GCF_000001635.18) 13 NC_000079.5 (21271828..21284726)
Chromosome 13 - NC_000079.6
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Transcript information: This gene has 7 transcripts
Gene: Trim27 ENSMUSG00000021326
Description tripartite motif-containing 27 [Source:MGI Symbol;Acc:MGI:97904] Gene Synonyms Rfp Location Chromosome 13: 21,179,445-21,194,724 forward strand. GRCm38:CM001006.2 About this gene This gene has 7 transcripts (splice variants), 152 orthologues, 73 paralogues, is a member of 1 Ensembl protein family and is associated with 12 phenotypes. Transcripts
Name Transcript ID bp Protein Translation ID Biotype CCDS UniProt Flags
Trim27- ENSMUST00000222544.1 4312 513aa ENSMUSP00000152730.1 Protein coding CCDS36607 Q62158 TSL:5 206 GENCODE basic APPRIS P1
Trim27- ENSMUST00000021761.12 4204 513aa ENSMUSP00000021761.5 Protein coding CCDS36607 Q62158 TSL:1 201 GENCODE basic APPRIS P1
Trim27- ENSMUST00000223065.1 2046 314aa ENSMUSP00000152179.1 Protein coding - A0A1Y7VKW0 TSL:1 207 GENCODE basic
Trim27- ENSMUST00000221464.1 1400 337aa ENSMUSP00000152230.1 Protein coding - A0A1Y7VJ13 TSL:5 205 GENCODE basic
Trim27- ENSMUST00000139287.3 683 61aa ENSMUSP00000115397.3 Nonsense mediated - F6U3T5 CDS 5' 204 decay incomplete TSL:5
Trim27- ENSMUST00000124052.2 1750 No - Retained intron - - TSL:NA 202 protein
Trim27- ENSMUST00000124794.1 439 No - Retained intron - - TSL:3 203 protein
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35.28 kb Forward strand 21.17Mb 21.18Mb 21.19Mb 21.20Mb Genes (Comprehensive set... Trim27-206 >protein coding
Trim27-201 >protein coding
Trim27-202 >retained intron Trim27-204 >nonsense mediated decay
Trim27-207 >protein coding
Trim27-205 >protein coding
Trim27-203 >retained intron
Contigs < AC138330.4
Genes < Gm47806-201lncRNA (Comprehensive set...
Regulatory Build
21.17Mb 21.18Mb 21.19Mb 21.20Mb Reverse strand 35.28 kb
Regulation Legend
CTCF Enhancer Open Chromatin Promoter Promoter Flank
Gene Legend Protein Coding
Ensembl protein coding
Non-Protein Coding
processed transcript RNA gene
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Transcript: ENSMUST00000021761
14.79 kb Forward strand
Trim27-201 >protein coding
ENSMUSP00000021... Low complexity (Seg) Coiled-coils (Ncoils) Superfamily SSF57850 SSF57845 Concanavalin A-like lectin/glucanase domain superfamily
SMART Zinc finger, RING-type SPRY-associated
B-box-type zinc finger SPRY domain Prints Zinc finger, B-box, chordata Butyrophylin-like, SPRY domain Pfam PF15227 B-box-type zinc finger SPRY domain
SPRY-associated PROSITE profiles Zinc finger, RING-type B30.2/SPRY domain
B-box-type zinc finger PROSITE patterns Zinc finger, RING-type, conserved site
PANTHER PTHR24103:SF362
PTHR24103 Gene3D Zinc finger, RING/FYVE/PHD-type 2.60.120.920
3.30.40.200 CDD cd16594 B-box-type zinc finger TRIM27, PRY/SPRY domain
All sequence SNPs/i... Sequence variants (dbSNP and all other sources)
Variant Legend
missense variant synonymous variant
Scale bar 0 60 120 180 240 300 360 420 513
We wish to acknowledge the following valuable scientific information resources: Ensembl, MGI, NCBI, UCSC.
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