https://www.alphaknockout.com
Mouse Fchsd1 Knockout Project (CRISPR/Cas9)
Objective: To create a Fchsd1 knockout Mouse model (C57BL/6J) by CRISPR/Cas-mediated genome engineering.
Strategy summary: The Fchsd1 gene (NCBI Reference Sequence: NM_175684 ; Ensembl: ENSMUSG00000038524 ) is located on Mouse chromosome 18. 20 exons are identified, with the ATG start codon in exon 1 and the TGA stop codon in exon 20 (Transcript: ENSMUST00000043437). Exon 1~18 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 1 starts from about 0.05% of the coding region. Exon 1~18 covers 94.23% of the coding region. The size of effective KO region: ~8478 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' 15
1 2 3 4 5 6 7 8 9 10 11 12 13 14 16 17 18 20
Legends Exon of mouse Fchsd1 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 start codon 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.
Overview of the Dot Plot (down) Window size: 15 bp
Forward Reverse Complement
Sequence 12
Note: The 2000 bp section downstream of Exon 18 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(26.9% 538) | C(22.55% 451) | T(26.65% 533) | G(23.9% 478)
Note: The 2000 bp section upstream of start codon 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(2000bp) | A(21.2% 424) | C(25.25% 505) | T(31.3% 626) | G(22.25% 445)
Note: The 2000 bp section downstream of Exon 18 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% chr18 - 37969727 37971726 2000 browser details YourSeq 160 740 1214 2000 85.0% chr3 - 96627254 96627482 229 browser details YourSeq 153 740 906 2000 96.4% chr1 + 135353340 135353701 362 browser details YourSeq 152 740 908 2000 95.9% chr9 - 88555337 88555506 170 browser details YourSeq 150 731 892 2000 95.1% chr3 - 142848486 142848646 161 browser details YourSeq 150 740 908 2000 95.2% chr16 + 33220496 33220664 169 browser details YourSeq 150 740 904 2000 95.8% chr1 + 74581960 74582141 182 browser details YourSeq 149 738 909 2000 95.8% chr1 + 86539783 86539955 173 browser details YourSeq 148 731 892 2000 94.5% chr17 - 35087560 35087720 161 browser details YourSeq 148 740 899 2000 95.6% chr6 + 122711960 122712118 159 browser details YourSeq 146 740 908 2000 94.6% chr10 - 43387188 43387357 170 browser details YourSeq 146 740 909 2000 94.0% chr10 - 4488034 4488215 182 browser details YourSeq 146 719 888 2000 95.6% chr8 + 71617013 71617188 176 browser details YourSeq 145 740 892 2000 97.4% chr4 - 46326578 46326730 153 browser details YourSeq 145 742 892 2000 98.1% chr17 - 42841435 42841585 151 browser details YourSeq 145 721 893 2000 93.5% chr11 - 6378781 6378964 184 browser details YourSeq 145 724 885 2000 96.2% chr6 + 99287305 99287478 174 browser details YourSeq 145 740 890 2000 98.1% chr5 + 122413720 122413870 151 browser details YourSeq 145 740 892 2000 97.4% chr3 + 94910925 94911077 153 browser details YourSeq 145 740 892 2000 97.4% chr12 + 101964664 101964816 153
Note: The 2000 bp section upstream of start codon 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 2000 1 2000 2000 100.0% chr18 - 37959247 37961246 2000 browser details YourSeq 96 406 896 2000 71.3% chr7 - 16350494 16350657 164 browser details YourSeq 91 413 573 2000 80.2% chr8 - 115129459 115129616 158 browser details YourSeq 91 406 531 2000 86.6% chr12 + 108540739 108541013 275 browser details YourSeq 86 406 527 2000 90.8% chr7 + 16802936 16803267 332 browser details YourSeq 85 394 537 2000 78.2% chr3 + 40487895 40488035 141 browser details YourSeq 84 406 533 2000 86.5% chr11 - 114898852 114898976 125 browser details YourSeq 83 402 526 2000 83.2% chr9 - 109221077 109221201 125 browser details YourSeq 83 406 516 2000 87.4% chr7 - 24657791 24657901 111 browser details YourSeq 83 394 516 2000 83.8% chr17 - 88198166 88198288 123 browser details YourSeq 83 401 531 2000 79.7% chr6 + 31312266 31312392 127 browser details YourSeq 82 406 564 2000 82.4% chr5 + 100252872 100253016 145 browser details YourSeq 82 406 586 2000 76.2% chr15 + 100243368 100243506 139 browser details YourSeq 81 402 516 2000 85.3% chr6 + 31378326 31378440 115 browser details YourSeq 79 416 567 2000 87.0% chr2 - 18651253 18651404 152 browser details YourSeq 78 406 517 2000 82.0% chr5 + 73044609 73044709 101 browser details YourSeq 78 393 516 2000 81.5% chr1 + 161517058 161517181 124 browser details YourSeq 77 406 531 2000 89.7% chr11 - 74878729 74879247 519 browser details YourSeq 76 406 531 2000 77.5% chr3 - 54594105 54594226 122 browser details YourSeq 75 406 520 2000 82.7% chr9 - 109554968 109555082 115
Note: The 2000 bp section downstream of Exon 18 is BLAT searched against the genome. No significant similarity is found.
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Gene and protein information: Fchsd1 FCH and double SH3 domains 1 [ Mus musculus (house mouse) ] Gene ID: 319262, updated on 12-Aug-2019
Gene summary
Official Symbol Fchsd1 provided by MGI Official Full Name FCH and double SH3 domains 1 provided by MGI Primary source MGI:MGI:2441771 See related Ensembl:ENSMUSG00000038524 Gene type protein coding RefSeq status PROVISIONAL Organism Mus musculus Lineage Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi; Mammalia; Eutheria; Euarchontoglires; Glires; Rodentia; Myomorpha; Muroidea; Muridae; Murinae; Mus; Mus Also known as NWK2; A030002D08Rik Expression Broad expression in CNS E18 (RPKM 14.4), cerebellum adult (RPKM 13.9) and 23 other tissues See more Orthologs human all
Genomic context
Location: 18; 18 B3 See Fchsd1 in Genome Data Viewer Exon count: 21
Annotation release Status Assembly Chr Location
108 current GRCm38.p6 (GCF_000001635.26) 18 NC_000084.6 (37957431..37969814, complement)
Build 37.2 previous assembly MGSCv37 (GCF_000001635.18) 18 NC_000084.5 (38117088..38129385, complement)
Chromosome 18 - NC_000084.6
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Transcript information: This gene has 10 transcripts
Gene: Fchsd1 ENSMUSG00000038524
Description FCH and double SH3 domains 1 [Source:MGI Symbol;Acc:MGI:2441771] Gene Synonyms A030002D08Rik Location Chromosome 18: 37,957,431-37,969,774 reverse strand. GRCm38:CM001011.2 About this gene This gene has 10 transcripts (splice variants), 178 orthologues, 1 paralogue and is a member of 1 Ensembl protein family. Transcripts
Name Transcript ID bp Protein Translation ID Biotype CCDS UniProt Flags
Fchsd1- ENSMUST00000043437.13 4270 688aa ENSMUSP00000047878.7 Protein coding CCDS37786 Q6PFY1 TSL:1 201 GENCODE basic APPRIS P1
Fchsd1- ENSMUST00000163131.7 454 137aa ENSMUSP00000133058.1 Protein coding - E9PVQ1 CDS 3' 202 incomplete TSL:3
Fchsd1- ENSMUST00000166148.7 369 108aa ENSMUSP00000129825.1 Protein coding - E9Q363 CDS 3' 205 incomplete TSL:2
Fchsd1- ENSMUST00000164222.1 419 57aa ENSMUSP00000128481.1 Nonsense mediated - E9QA76 TSL:2 203 decay
Fchsd1- ENSMUST00000165643.1 2918 No - Retained intron - - TSL:1 204 protein
Fchsd1- ENSMUST00000166973.1 690 No - Retained intron - - TSL:5 207 protein
Fchsd1- ENSMUST00000168136.7 526 No - Retained intron - - TSL:3 208 protein
Fchsd1- ENSMUST00000166727.1 422 No - Retained intron - - TSL:3 206 protein
Fchsd1- ENSMUST00000171253.1 357 No - Retained intron - - TSL:3 210 protein
Fchsd1- ENSMUST00000168816.1 350 No - Retained intron - - TSL:2 209 protein
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32.34 kb Forward strand
37.95Mb 37.96Mb 37.97Mb Genes Rell2-203 >nonsense mediated decay (Comprehensive set...
Rell2-211 >protein coding
Rell2-207 >nonsense mediated decay
Rell2-210 >protein coding
Rell2-201 >protein coding
Rell2-212 >protein coding
Rell2-208 >protein coding
Rell2-204 >protein coding
Rell2-202 >protein coding
Rell2-209 >protein coding
Rell2-213 >lncRNA
Rell2-215 >lncRNA
Rell2-205 >retained intron
Rell2-206 >retained intron
Rell2-214 >retained intron
Contigs AC129315.3 > Genes (Comprehensive set... < Hdac3-201protein coding < Fchsd1-204retained intron < Fchsd1-206retained intro < Hdac3-206nonsense mediated decay < Fchsd1-201protein coding < Arap3-201protein coding < Hdac3-205nonsense mediated decay < Fchsd1-205protein coding < Arap3-206retained intron < Fchsd1-207retained intron < Arap3-204protein coding < Fchsd1-202protein coding < Mir6981-201miRNA < Fchsd1-208retained intron < Fchsd1-203nonsense mediated decay < Fchsd1-210retained intron Regulatory Build 37.95Mb 37.96Mb 37.97Mb Reverse strand 32.34 kb Regulation Legend CTCF Enhancer Promoter Promoter Flank Gene Legend Protein Coding merged Ensembl/Havana Ensembl protein coding Non-Protein Coding processed transcript RNA gene Page 8 of 10 https://www.alphaknockout.com Page 9 of 10 https://www.alphaknockout.com Transcript: ENSMUST00000043437 < Fchsd1-201protein coding Reverse strand 12.34 kb ENSMUSP00000047... MobiDB lite Low complexity (Seg) Coiled-coils (Ncoils) Superfamily AH/BAR domain superfamily SH3-like domain superfamily SMART FCH domain SH3 domain Pfam FCH domain SH3 domain SH3 domain PROSITE profiles F-BAR domain SH3 domain PANTHER F-BAR and double SH3 domains protein 1 PTHR15735 Gene3D AH/BAR domain superfamily 2.30.30.40 CDD FCHSD, SH3 domain 1 All sequence SNPs/i... Sequence variants (dbSNP and all other sources) Variant Legend missense variant synonymous variant Scale bar 0 60 120 180 240 300 360 420 480 540 600 688 We wish to acknowledge the following valuable scientific information resources: Ensembl, MGI, NCBI, UCSC. Page 10 of 10