https://www.alphaknockout.com
Mouse Kirrel2 Knockout Project (CRISPR/Cas9)
Objective: To create a Kirrel2 knockout Mouse model (C57BL/6J) by CRISPR/Cas-mediated genome engineering.
Strategy summary: The Kirrel2 gene (NCBI Reference Sequence: NM_172898 ; Ensembl: ENSMUSG00000036915 ) is located on Mouse chromosome 7. 15 exons are identified, with the ATG start codon in exon 1 and the TGA stop codon in exon 15 (Transcript: ENSMUST00000045817). Exon 3~15 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: Mice homozygous for a knock-out allele exhibit defects in glomeruli formation in the posterior region of the accessory olfactory bulb.
Exon 3 starts from about 9.67% of the coding region. Exon 3~15 covers 90.38% of the coding region. The size of effective KO region: ~7491 bp. The KO region does not have any other known gene.
Page 1 of 8 https://www.alphaknockout.com
Overview of the Targeting Strategy
Wildtype allele 5' gRNA region gRNA region 3'
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
Legends Exon of mouse Kirrel2 Knockout region
Page 2 of 8 https://www.alphaknockout.com
Overview of the Dot Plot (up) Window size: 15 bp
Forward Reverse Complement
Sequence 12
Note: The 2000 bp section upstream of Exon 3 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 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.
Page 3 of 8 https://www.alphaknockout.com
Overview of the GC Content Distribution (up) Window size: 300 bp
Sequence 12
Summary: Full Length(2000bp) | A(19.95% 399) | C(27.3% 546) | T(26.75% 535) | G(26.0% 520)
Note: The 2000 bp section upstream of Exon 3 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(28.3% 566) | C(22.45% 449) | T(24.8% 496) | G(24.45% 489)
Note: The 2000 bp section downstream of stop 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.
Page 4 of 8 https://www.alphaknockout.com
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 - 30455615 30457614 2000 browser details YourSeq 93 1528 1855 2000 85.4% chr11 - 23253209 23627850 374642 browser details YourSeq 54 1508 1859 2000 90.8% chr19 - 53509131 53509491 361 browser details YourSeq 54 1492 1802 2000 69.6% chr16 + 11101106 11101295 190 browser details YourSeq 49 1531 1848 2000 86.6% chr1 - 161030625 161031063 439 browser details YourSeq 49 1494 1559 2000 88.9% chr8 + 72344573 72344642 70 browser details YourSeq 47 1729 1848 2000 86.5% chr16 - 72674866 72674984 119 browser details YourSeq 46 1720 1848 2000 94.3% chr8 + 94651641 94651781 141 browser details YourSeq 46 1729 1848 2000 87.5% chr11 + 20503590 20503708 119 browser details YourSeq 45 1729 1848 2000 80.9% chr10 - 91055532 91055653 122 browser details YourSeq 44 1731 1848 2000 87.1% chr16 - 71798180 71798296 117 browser details YourSeq 43 1508 1559 2000 92.2% chr9 + 67844459 67844511 53 browser details YourSeq 42 1720 1801 2000 93.7% chr3 - 108446417 108446510 94 browser details YourSeq 42 1504 1555 2000 90.4% chr11 - 116200928 116200979 52 browser details YourSeq 42 1505 1564 2000 90.4% chr11 + 69313944 69314004 61 browser details YourSeq 41 235 307 2000 79.2% chr1 - 171009109 171009174 66 browser details YourSeq 41 1787 1858 2000 77.1% chr11 + 83921549 83921617 69 browser details YourSeq 40 1723 1848 2000 83.7% chr9 - 108418109 108418232 124 browser details YourSeq 39 248 294 2000 90.5% chr8 - 128313675 128313719 45 browser details YourSeq 39 249 302 2000 83.8% chr2 + 142188478 142188526 49
Note: The 2000 bp section upstream of Exon 3 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 - 30446122 30448121 2000 browser details YourSeq 418 601 1316 2000 90.5% chr18 - 74209489 74210078 590 browser details YourSeq 414 600 1096 2000 93.0% chr1 - 4921631 4922129 499 browser details YourSeq 409 601 1095 2000 92.4% chr17 + 50647879 50648368 490 browser details YourSeq 397 600 1092 2000 92.6% chr18 - 34191125 34191612 488 browser details YourSeq 396 601 1096 2000 91.8% chr11 - 19814339 19814828 490 browser details YourSeq 394 600 1106 2000 89.9% chr11 + 63200212 63200711 500 browser details YourSeq 391 601 1095 2000 90.0% chr2 + 17098958 17099449 492 browser details YourSeq 390 601 1095 2000 91.0% chr2 - 159433761 159434250 490 browser details YourSeq 388 601 1092 2000 91.2% chr1 - 43333312 43333799 488 browser details YourSeq 387 601 1095 2000 91.1% chr13 - 32561175 32561663 489 browser details YourSeq 387 601 1093 2000 90.2% chr12 + 16406371 16406858 488 browser details YourSeq 386 600 1095 2000 90.1% chr7 + 18833321 18833811 491 browser details YourSeq 385 601 1095 2000 90.6% chr3 - 33575238 33575727 490 browser details YourSeq 385 601 1096 2000 89.4% chr9 + 20423657 20424149 493 browser details YourSeq 382 604 1095 2000 89.6% chr10 - 66855558 66856029 472 browser details YourSeq 381 601 1097 2000 90.0% chr8 - 129267161 129267642 482 browser details YourSeq 379 601 1095 2000 89.6% chr9 + 89469508 89469997 490 browser details YourSeq 378 601 1095 2000 91.1% chr16 - 8952619 8953109 491 browser details YourSeq 378 601 1096 2000 90.4% chr6 + 107058064 107058552 489
Note: The 2000 bp section downstream of stop codon is BLAT searched against the genome. No significant similarity is found.
Page 5 of 8 https://www.alphaknockout.com
Gene and protein information: Kirrel2 kirre like nephrin family adhesion molecule 2 [ Mus musculus (house mouse) ] Gene ID: 243911, updated on 22-Oct-2019
Gene summary
Official Symbol Kirrel2 provided by MGI Official Full Name kirre like nephrin family adhesion molecule 2 provided by MGI Primary source MGI:MGI:2442334 See related Ensembl:ENSMUSG00000036915 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 NLG1; NEPH3; C330019F22Rik Expression Biased expression in cortex adult (RPKM 1.4), CNS E11.5 (RPKM 1.2) and 6 other tissues See more Orthologs human all
Genomic context
Location: 7; 7 B1 See Kirrel2 in Genome Data Viewer Exon count: 15
Annotation release Status Assembly Chr Location
108 current GRCm38.p6 (GCF_000001635.26) 7 NC_000073.6 (30447766..30457515, complement)
Build 37.2 previous assembly MGSCv37 (GCF_000001635.18) 7 NC_000073.5 (31232785..31242534, complement)
Chromosome 7 - NC_000073.6
Page 6 of 8 https://www.alphaknockout.com
Transcript information: This gene has 5 transcripts
Gene: Kirrel2 ENSMUSG00000036915
Description kirre like nephrin family adhesion molecule 2 [Source:MGI Symbol;Acc:MGI:2442334] Gene Synonyms C330019F22Rik, NEPH3 Location Chromosome 7: 30,447,534-30,457,690 reverse strand. GRCm38:CM001000.2 About this gene This gene has 5 transcripts (splice variants), 169 orthologues, 3 paralogues, is a member of 1 Ensembl protein family and is associated with 1 phenotype. Transcripts
Name Transcript ID bp Protein Translation ID Biotype CCDS UniProt Flags
Kirrel2- ENSMUST00000045817.13 3352 700aa ENSMUSP00000039395.7 Protein CCDS39883 A0A4X8 TSL:1 201 coding Q7TSU7 GENCODE basic APPRIS P1
Kirrel2- ENSMUST00000169893.2 930 310aa ENSMUSP00000131161.2 Protein - A0A171EBL3 CDS 5' and 3' 204 coding incomplete TSL:1
Kirrel2- ENSMUST00000170152.7 575 175aa ENSMUSP00000132652.1 Protein - F6WZB3 CDS 5' incomplete 205 coding TSL:3
Kirrel2- ENSMUST00000140565.1 3901 No - Retained - - TSL:1 203 protein intron
Kirrel2- ENSMUST00000117923.1 343 No - lncRNA - - TSL:3 202 protein
30.16 kb Forward strand 30.44Mb 30.45Mb 30.46Mb Genes Nphs1-203 >protein coding (Comprehensive set...
Nphs1-201 >protein coding
Contigs AC149067.3 > < AC167970.4 Genes (Comprehensive set... < Aplp1-201protein coding < Kirrel2-203retained intron < Nphs1os-201lncRNA
< Aplp1-204retained intron < Kirrel2-201protein coding
< Aplp1-202retained intron < Kirrel2-205protein coding < Kirrel2-204protein coding
< Aplp1-205retained intron < Kirrel2-202lncRNA
Regulatory Build
30.44Mb 30.45Mb 30.46Mb Reverse strand 30.16 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
Page 7 of 8 https://www.alphaknockout.com
Transcript: ENSMUST00000045817
< Kirrel2-201protein coding
Reverse strand 9.98 kb
ENSMUSP00000039... Transmembrane heli... MobiDB lite Low complexity (Seg) Cleavage site (Sign... Superfamily Immunoglobulin-like domain superfamily SMART Immunoglobulin subtype
Immunoglobulin subtype 2 Pfam Immunoglobulin I-set CD80-like, immunoglobulin C2-set
PF13927 PROSITE profiles Immunoglobulin-like domain PANTHER PTHR11640
PTHR11640:SF51 Gene3D Immunoglobulin-like fold
All sequence SNPs/i... Sequence variants (dbSNP and all other sources)
Variant Legend missense variant splice region variant synonymous variant
Scale bar 0 60 120 180 240 300 360 420 480 540 600 700
We wish to acknowledge the following valuable scientific information resources: Ensembl, MGI, NCBI, UCSC.
Page 8 of 8