https://www.alphaknockout.com
Mouse Calm3 Knockout Project (CRISPR/Cas9)
Objective: To create a Calm3 knockout Mouse model (C57BL/6J) by CRISPR/Cas-mediated genome engineering.
Strategy summary: The Calm3 gene (NCBI Reference Sequence: NM_007590 ; Ensembl: ENSMUSG00000019370 ) is located on Mouse chromosome 7. 6 exons are identified, with the ATG start codon in exon 1 and the TGA stop codon in exon 6 (Transcript: ENSMUST00000019514). Exon 1~6 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 0.22% of the coding region. Exon 1~6 covers 100.0% of the coding region. The size of effective KO region: ~6891 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 6
Legends Exon of mouse Calm3 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 stop 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.
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Overview of the GC Content Distribution (up) Window size: 300 bp
Sequence 12
Summary: Full Length(2000bp) | A(24.5% 490) | C(26.35% 527) | T(23.7% 474) | G(25.45% 509)
Note: The 2000 bp section upstream of start codon is analyzed to determine the GC content. Significant high GC-content regions are found. The gRNA site is selected outside of these high GC-content regions.
Overview of the GC Content Distribution (down) Window size: 300 bp
Sequence 12
Summary: Full Length(2000bp) | A(18.8% 376) | C(28.5% 570) | T(24.1% 482) | G(28.6% 572)
Note: The 2000 bp section downstream of stop codon is analyzed to determine the GC content. Significant high GC-content regions are found. The gRNA site is selected outside of these high GC-content regions.
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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% chr7 - 16923844 16925843 2000 browser details YourSeq 107 22 241 2000 92.2% chr17 - 48764639 48765114 476 browser details YourSeq 99 1 124 2000 91.9% chr5 - 110853122 110853243 122 browser details YourSeq 89 1 212 2000 93.3% chr3 - 86268930 86269354 425 browser details YourSeq 89 2 450 2000 76.0% chr15 - 78898190 78898321 132 browser details YourSeq 83 1 102 2000 92.0% chr6 - 99542889 99542992 104 browser details YourSeq 78 3 209 2000 91.5% chr5 + 53459865 53460107 243 browser details YourSeq 75 1 85 2000 94.2% chr18 - 34124216 34124300 85 browser details YourSeq 74 1 90 2000 91.2% chr10 + 41355633 41355722 90 browser details YourSeq 73 2 80 2000 96.3% chr12 - 87035636 87035714 79 browser details YourSeq 73 147 242 2000 94.2% chr12 - 11397650 11397767 118 browser details YourSeq 72 4 85 2000 94.0% chr16 - 34882096 34882177 82 browser details YourSeq 72 147 451 2000 94.0% chr13 + 59849536 59849985 450 browser details YourSeq 71 1 85 2000 91.8% chr5 - 65959913 65959997 85 browser details YourSeq 71 1 77 2000 96.2% chr2 + 165130306 165130382 77 browser details YourSeq 70 147 240 2000 93.8% chr2 - 93664012 93664128 117 browser details YourSeq 70 147 241 2000 88.9% chr8 + 40470416 40470511 96 browser details YourSeq 69 121 210 2000 88.8% chr4 - 127301023 127301112 90 browser details YourSeq 69 1 123 2000 92.3% chr17 - 46871480 46871601 122 browser details YourSeq 67 29 124 2000 85.3% chr2 - 69978895 69978989 95
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% chr7 - 16914951 16916950 2000 browser details YourSeq 134 1 142 2000 97.2% chr6 - 96032860 96033001 142 browser details YourSeq 29 1957 1986 2000 100.0% chr7 + 24962412 24962442 31 browser details YourSeq 28 563 593 2000 96.7% chr12 + 74445529 74445566 38 browser details YourSeq 26 691 717 2000 100.0% chr8 + 90592281 90592310 30 browser details YourSeq 26 567 595 2000 96.5% chr2 + 17857314 17857638 325 browser details YourSeq 25 1945 1971 2000 96.3% chr9 - 7685440 7685466 27 browser details YourSeq 24 1436 1460 2000 100.0% chr17 + 17163826 17163851 26 browser details YourSeq 23 279 301 2000 100.0% chr13 - 12101465 12101487 23 browser details YourSeq 23 565 588 2000 100.0% chr4 + 57913833 57913858 26 browser details YourSeq 23 1463 1485 2000 100.0% chr16 + 23987374 23987396 23 browser details YourSeq 22 565 587 2000 100.0% chr4 - 20326333 20326357 25 browser details YourSeq 21 566 586 2000 100.0% chr12 - 46694476 46694496 21 browser details YourSeq 21 566 586 2000 100.0% chr1 + 79776696 79776716 21
Note: The 2000 bp section downstream of stop codon is BLAT searched against the genome. No significant similarity is found.
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Gene and protein information: Calm3 calmodulin 3 [ Mus musculus (house mouse) ] Gene ID: 12315, updated on 10-Oct-2019
Gene summary
Official Symbol Calm3 provided by MGI Official Full Name calmodulin 3 provided by MGI Primary source MGI:MGI:103249 See related Ensembl:ENSMUSG00000019370 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 CaMA; Cam3; Camc; R75142 Expression Ubiquitous expression in cortex adult (RPKM 496.4), frontal lobe adult (RPKM 463.0) and 28 other tissues See more Orthologs human all
Genomic context
Location: 7 A2; 7 9.15 cM See Calm3 in Genome Data Viewer Exon count: 6
Annotation release Status Assembly Chr Location
108 current GRCm38.p6 (GCF_000001635.26) 7 NC_000073.6 (16915379..16924032, complement)
Build 37.2 previous assembly MGSCv37 (GCF_000001635.18) 7 NC_000073.5 (17500728..17509381, complement)
Chromosome 7 - NC_000073.6
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Transcript information: This gene has 4 transcripts
Gene: Calm3 ENSMUSG00000019370
Description calmodulin 3 [Source:MGI Symbol;Acc:MGI:103249] Location Chromosome 7: 16,915,379-16,924,114 reverse strand. GRCm38:CM001000.2 About this gene This gene has 4 transcripts (splice variants), 102 orthologues, 22 paralogues and is a member of 1 Ensembl protein family. Transcripts
Name Transcript ID bp Protein Translation ID Biotype CCDS UniProt Flags
Calm3- ENSMUST00000019514.9 2290 149aa ENSMUSP00000019514.9 Protein coding CCDS39789 P0DP26 P0DP27 TSL:1 201 P0DP28 GENCODE basic APPRIS P1
Calm3- ENSMUST00000173139.1 490 65aa ENSMUSP00000134395.1 Protein coding - G3UZ90 CDS 5' 203 incomplete TSL:3
Calm3- ENSMUST00000172594.1 648 39aa ENSMUSP00000133559.1 Nonsense mediated - G3UX57 CDS 5' 202 decay incomplete TSL:5
Calm3- ENSMUST00000173557.1 482 No - lncRNA - - TSL:5 204 protein
28.74 kb Forward strand 16.91Mb 16.92Mb 16.93Mb Genes Ptgir-201 >protein coding (Comprehensive set...
Ptgir-202 >protein coding
Contigs < AC148981.7 < AC165955.3 Genes (Comprehensive set... < Calm3-201protein coding
< Calm3-202nonsense mediated decay
< Calm3-203protein coding
< Calm3-204lncRNA
Regulatory Build
16.91Mb 16.92Mb 16.93Mb Reverse strand 28.74 kb
Regulation Legend CTCF Enhancer Open Chromatin Promoter Promoter Flank
Gene Legend Protein Coding
merged Ensembl/Havana Ensembl protein coding
Non-Protein Coding
processed transcript RNA gene
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Transcript: ENSMUST00000019514
< Calm3-201protein coding
Reverse strand 8.74 kb
ENSMUSP00000019... PDB-ENSP mappings Superfamily EF-hand domain pair
SMART EF-hand domain Prints PR00450 Pfam EF-hand domain PROSITE profiles EF-hand domain PROSITE patterns EF-Hand 1, calcium-binding site PANTHER Calmodulin
PTHR23050 Gene3D 1.10.238.10 CDD EF-hand domain
All sequence SNPs/i... Sequence variants (dbSNP and all other sources)
Variant Legend
synonymous variant
Scale bar 0 20 40 60 80 100 120 149
We wish to acknowledge the following valuable scientific information resources: Ensembl, MGI, NCBI, UCSC.
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