https://www.alphaknockout.com
Mouse Magi2 Conditional Knockout Project (CRISPR/Cas9)
Objective: To create a Magi2 conditional knockout Mouse model (C57BL/6J) by CRISPR/Cas-mediated genome engineering.
Strategy summary: The Magi2 gene (NCBI Reference Sequence: NM_001170746 ; Ensembl: ENSMUSG00000040003 ) is located on Mouse chromosome 5. 23 exons are identified, with the ATG start codon in exon 1 and the TAG stop codon in exon 23 (Transcript: ENSMUST00000088516). Exon 12 will be selected as conditional knockout region (cKO region). Deletion of this region should result in the loss of function of the Mouse Magi2 gene. To engineer the targeting vector, homologous arms and cKO region will be generated by PCR using BAC clone RP23-425D14 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: Homozygotes for a null allele show neonatal death and hippocampal neurons with altered dendritic spine morphology. Homozygotes for a different null allele die neonatally due to anuria and podocyte anomalies. Mice lacking all three isoforms develop proteinuria, podocytopathy and die of renal failure.
Exon 12 starts from about 54.3% of the coding region. The knockout of Exon 12 will result in frameshift of the gene. The size of intron 11 for 5'-loxP site insertion: 6016 bp, and the size of intron 12 for 3'-loxP site insertion: 9058 bp. The size of effective cKO region: ~690 bp. The cKO region does not have any other known gene.
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Overview of the Targeting Strategy
Wildtype allele gRNA region 5' gRNA region 3'
1 12 23 Targeting vector
Targeted allele
Constitutive KO allele (After Cre recombination)
Legends Exon of mouse Magi2 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. 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(7190bp) | A(29.6% 2128) | C(21.29% 1531) | T(27.69% 1991) | G(21.42% 1540)
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% chr5 + 20531088 20534087 3000 browser details YourSeq 195 1252 1729 3000 87.5% chr8 - 78673395 78673755 361 browser details YourSeq 183 1252 1716 3000 86.0% chr3 - 35702703 35702923 221 browser details YourSeq 181 1253 1730 3000 85.2% chr8 - 65289317 65289528 212 browser details YourSeq 180 1252 1453 3000 95.0% chr1 - 58466182 58466387 206 browser details YourSeq 177 1252 1455 3000 93.7% chr2 - 63879656 63879868 213 browser details YourSeq 177 1252 1457 3000 94.1% chr10 - 98055730 98055937 208 browser details YourSeq 176 1252 1729 3000 83.8% chr14 + 32488515 32488725 211 browser details YourSeq 176 1252 1713 3000 84.0% chr11 + 106260319 106260540 222 browser details YourSeq 175 1252 1453 3000 93.6% chrX - 106766309 106766512 204 browser details YourSeq 175 1245 1452 3000 92.5% chr5 + 45054345 45054550 206 browser details YourSeq 175 1252 1450 3000 94.0% chr12 + 119047475 119047673 199 browser details YourSeq 174 1252 1448 3000 94.4% chr9 - 90329376 90329580 205 browser details YourSeq 174 1252 1447 3000 93.4% chr15 - 13427968 13428162 195 browser details YourSeq 174 1252 1449 3000 94.0% chr10 - 123246402 123246599 198 browser details YourSeq 174 1252 1447 3000 94.4% chr1 - 135620698 135620893 196 browser details YourSeq 174 1252 1451 3000 94.5% chr5 + 102235184 102235394 211 browser details YourSeq 174 1252 1453 3000 93.6% chr17 + 65074262 65074470 209 browser details YourSeq 173 1252 1448 3000 92.9% chr12 + 78484714 78484909 196 browser details YourSeq 173 1252 1450 3000 93.5% chr10 + 74787643 74787841 199
Note: The 3000 bp section upstream of Exon 12 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% chr5 + 20534778 20537777 3000 browser details YourSeq 151 577 776 3000 93.1% chr4 + 88741218 88741416 199 browser details YourSeq 149 592 773 3000 91.3% chr16 + 3918223 3918384 162 browser details YourSeq 148 592 773 3000 91.2% chr5 - 29748954 29749114 161 browser details YourSeq 147 582 771 3000 94.7% chr14 + 121449423 121449659 237 browser details YourSeq 147 592 774 3000 90.7% chr13 + 94361453 94361614 162 browser details YourSeq 145 590 774 3000 89.8% chr2 + 60152252 60152407 156 browser details YourSeq 144 483 773 3000 94.0% chr18 - 60822171 60822600 430 browser details YourSeq 142 592 775 3000 96.2% chr5 - 32876046 32876352 307 browser details YourSeq 142 593 772 3000 90.5% chr11 + 5850363 5850527 165 browser details YourSeq 141 592 776 3000 92.0% chr10 - 80378015 80378197 183 browser details YourSeq 141 592 778 3000 95.5% chr1 + 191835920 191836118 199 browser details YourSeq 140 593 773 3000 90.0% chrX - 41134876 41135028 153 browser details YourSeq 140 316 721 3000 85.2% chr3 - 115984008 115984225 218 browser details YourSeq 140 579 772 3000 89.2% chr1 + 46341078 46341267 190 browser details YourSeq 139 590 772 3000 90.1% chr4 - 34641647 34641809 163 browser details YourSeq 139 593 773 3000 88.8% chr2 - 94015941 94016091 151 browser details YourSeq 139 593 773 3000 89.4% chr16 - 14375616 14375766 151 browser details YourSeq 139 592 773 3000 88.2% chr14 - 105058578 105058739 162 browser details YourSeq 138 592 773 3000 88.8% chr4 - 108591073 108591224 152
Note: The 3000 bp section downstream of Exon 12 is BLAT searched against the genome. No significant similarity is found.
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Gene and protein information: Magi2 membrane associated guanylate kinase, WW and PDZ domain containing 2 [ Mus musculus (house mouse) ] Gene ID: 50791, updated on 12-Aug-2019
Gene summary
Official Symbol Magi2 provided by MGI Official Full Name membrane associated guanylate kinase, WW and PDZ domain containing 2 provided by MGI Primary source MGI:MGI:1354953 See related Ensembl:ENSMUSG00000040003 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 AIP-1; Acvri1; Magi-2; S-SCAM; Acvrip1; Acvrinp1; mKIAA0705 Expression Broad expression in cortex adult (RPKM 7.1), frontal lobe adult (RPKM 6.2) and 15 other tissues See more Orthologs human all
Genomic context
Location: 5; 5 A3 See Magi2 in Genome Data Viewer
Exon count: 28
Annotation release Status Assembly Chr Location
108 current GRCm38.p6 (GCF_000001635.26) 5 NC_000071.6 (19219387..20704792)
Build 37.2 previous assembly MGSCv37 (GCF_000001635.18) 5 NC_000071.5 (18732864..20210610)
Chromosome 5 - NC_000071.6
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Transcript information: This gene has 15 transcripts
Gene: Magi2 ENSMUSG00000040003
Description membrane associated guanylate kinase, WW and PDZ domain containing 2 [Source:MGI Symbol;Acc:MGI:1354953] Gene Synonyms Acvrinp1, Magi-2, S-SCAM Location Chromosome 5: 19,227,036-20,704,792 forward strand. GRCm38:CM000998.2 About this gene This gene has 15 transcripts (splice variants), 284 orthologues, 6 paralogues, is a member of 1 Ensembl protein family and is associated with 34 phenotypes. Transcripts
Name Transcript ID bp Protein Translation ID Biotype CCDS UniProt Flags
Magi2- ENSMUST00000101558.9 6530 1098aa ENSMUSP00000099094.3 Protein coding CCDS51425 Q9WVQ1 TSL:1 202 GENCODE basic
Magi2- ENSMUST00000088516.9 4843 1275aa ENSMUSP00000085872.3 Protein coding CCDS51424 Q9WVQ1 TSL:5 201 GENCODE basic APPRIS P2
Magi2- ENSMUST00000115267.6 4478 1112aa ENSMUSP00000110922.2 Protein coding CCDS39019 Q9WVQ1 TSL:1 203 GENCODE basic
Magi2- ENSMUST00000197443.4 6578 1400aa ENSMUSP00000142764.1 Protein coding - A0A0G2JEG6 TSL:5 206 GENCODE basic APPRIS ALT2
Magi2- ENSMUST00000197354.4 4785 1414aa ENSMUSP00000142576.1 Protein coding - A0A0G2JE00 TSL:5 205 GENCODE basic APPRIS ALT2
Magi2- ENSMUST00000208219.1 3266 837aa ENSMUSP00000146458.1 Protein coding - A0A140LHL1 TSL:5 215 GENCODE basic
Magi2- ENSMUST00000197553.4 3264 853aa ENSMUSP00000146769.1 Protein coding - Q9WVQ1 TSL:1 207 GENCODE basic
Magi2- ENSMUST00000199514.1 899 188aa ENSMUSP00000143578.1 Protein coding - A0A0G2JGI5 CDS 5' incomplete 211 TSL:5
Magi2- ENSMUST00000207284.1 714 238aa ENSMUSP00000146348.1 Protein coding - A0A140LHB5 CDS 5' and 3' 214 incomplete TSL:5
Magi2- ENSMUST00000200443.1 417 38aa ENSMUSP00000142581.1 Protein coding - A0A0G2JE05 CDS 3' incomplete 212 TSL:5
Magi2- ENSMUST00000199384.1 3125 No - Retained - - TSL:NA 210 protein intron
Magi2- ENSMUST00000197822.4 853 No - lncRNA - - TSL:3 208 protein
Magi2- ENSMUST00000198908.1 831 No - lncRNA - - TSL:5 209 protein
Magi2- ENSMUST00000197307.1 826 No - lncRNA - - TSL:5 204 protein
Magi2- ENSMUST00000200518.1 756 No - lncRNA - - TSL:5 213 protein
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1.50 Mb Forward strand 19.4Mb 19.6Mb 19.8Mb 20.0Mb 20.2Mb 20.4Mb 20.6Mb Genes Magi2-205 >protein coding (Comprehensive set...
Magi2-213 >lncRNA Magi2-202 >protein coding
Magi2-201 >protein coding
Magi2-206 >protein coding
Magi2-210 >retained intron Magi2-203 >protein coding
Magi2-212 >protein coding Magi2-208 >lncRNA Magi2-214 >protein coding
Gm23570-201 >snRNA Gm3544-201 >processed pseudogene
Magi2-215 >protein coding
Magi2-207 >protein coding
Gm22755-201 >snRNA Magi2-211 >protein coding
Magi2-209 >lncRNA
Gm29254-201 >lncRNA
Gm29254-202 >lncRNA
Magi2-204 >lncRNA
Contigs AC113596.8 > < AC158141.8
Genes < 4921504A21Rik-201lncRNA < Gm21009-201processed pseudogene < Gm25761-201snoRNA (Comprehensive set...
< 4921504A21Rik-202lncRNA < Gm25335-201snRNA < Gm43680-201processed pseudogene
Regulatory Build
19.4Mb 19.6Mb 19.8Mb 20.0Mb 20.2Mb 20.4Mb 20.6Mb Reverse strand 1.50 Mb
Regulation Legend CTCF Enhancer Open Chromatin Promoter Promoter Flank Transcription Factor Binding Site
Gene Legend Protein Coding
Ensembl protein coding merged Ensembl/Havana
Non-Protein Coding
RNA gene processed transcript pseudogene
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Transcript: ENSMUST00000088516
1.48 Mb Forward strand
Magi2-201 >protein coding
ENSMUSP00000085... MobiDB lite Low complexity (Seg) Superfamily PDZ superfamily
WW domain superfamily
P-loop containing nucleoside triphosphate hydrolase SMART Guanylate kinase/L-type calcium channel beta subunit
PDZ domain
WW domain Pfam PF16663 WW domain PDZ domain 6
PDZ domain
Guanylate kinase/L-type calcium channel beta subunit PROSITE profiles Guanylate kinase-like domain
PDZ domain
WW domain PROSITE patterns WW domain
Guanylate kinase, conserved site PANTHER Membrane-associated guanylate kinase, WW and PDZ domain-containing protein 2
PTHR10316 Gene3D 3.30.63.10 2.20.70.10
2.30.42.10 CDD cd00992
WW domain
All sequence SNPs/i... Sequence variants (dbSNP and all other sources)
Variant Legend start lost missense variant splice region variant synonymous variant
Scale bar 0 200 400 600 800 1000 1275
We wish to acknowledge the following valuable scientific information resources: Ensembl, MGI, NCBI, UCSC.
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