https://www.alphaknockout.com
Mouse Pcdh7 Knockout Project (CRISPR/Cas9)
Objective: To create a Pcdh7 knockout Mouse model (C57BL/6J) by CRISPR/Cas-mediated genome engineering.
Strategy summary: The Pcdh7 gene (NCBI Reference Sequence: NM_001122758 ; Ensembl: ENSMUSG00000029108 ) is located on Mouse chromosome 5. 3 exons are identified, with the ATG start codon in exon 1 and the TAG stop codon in exon 3 (Transcript: ENSMUST00000191837). Exon 1 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 the coding region. Exon 1 covers 84.3% of the coding region. The size of effective KO region: ~3174 bp. The KO region does not have any other known gene.
Page 1 of 9 https://www.alphaknockout.com
Overview of the Targeting Strategy
Wildtype allele 5' gRNA region gRNA region 3'
1 3
Legends Exon of mouse Pcdh7 Knockout region
Page 2 of 9 https://www.alphaknockout.com
Overview of the Dot Plot (up) Window size: 15 bp
Forward Reverse Complement
Sequence 12
Note: The 2000 bp section of Exon 1 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 of Exon 1 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.
Page 3 of 9 https://www.alphaknockout.com
Overview of the GC Content Distribution (up) Window size: 300 bp
Sequence 12
Summary: Full Length(2000bp) | A(22.05% 441) | C(27.95% 559) | T(18.6% 372) | G(31.4% 628)
Note: The 2000 bp section of Exon 1 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(29.35% 587) | C(24.4% 488) | T(22.1% 442) | G(24.15% 483)
Note: The 2000 bp section of Exon 1 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 9 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% chr5 + 57719107 57721106 2000 browser details YourSeq 27 1513 1542 2000 96.6% chr12 - 105875862 105876013 152 browser details YourSeq 26 1509 1537 2000 96.5% chr1 - 63996257 63996288 32 browser details YourSeq 25 1571 1597 2000 96.3% chr18 + 37727216 37727242 27 browser details YourSeq 24 1963 1994 2000 74.1% chr10 - 81201706 81201732 27 browser details YourSeq 23 354 377 2000 100.0% chr1 + 34643101 34643136 36 browser details YourSeq 20 1817 1838 2000 95.5% chr12 + 79717096 79717117 22 browser details YourSeq 20 410 429 2000 100.0% chr10 + 74326124 74326143 20
Note: The 2000 bp section of Exon 1 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% chr5 + 57720279 57722278 2000 browser details YourSeq 35 399 470 2000 94.9% chr18 + 37727216 37727287 72 browser details YourSeq 26 337 365 2000 96.5% chr1 - 63996257 63996288 32 browser details YourSeq 25 965 989 2000 100.0% chr18 + 37690593 37690617 25 browser details YourSeq 23 1564 1587 2000 100.0% chr1 + 29846190 29846216 27 browser details YourSeq 20 694 715 2000 95.5% chr10 - 13986370 13986391 22
Note: The 2000 bp section of Exon 1 is BLAT searched against the genome. No significant similarity is found.
Page 5 of 9 https://www.alphaknockout.com
Gene and protein information: Pcdh7 protocadherin 7 [ Mus musculus (house mouse) ] Gene ID: 54216, updated on 12-Aug-2019
Gene summary
Official Symbol Pcdh7 provided by MGI Official Full Name protocadherin 7 provided by MGI Primary source MGI:MGI:1860487 See related Ensembl:ENSMUSG00000029108 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 Expression Broad expression in bladder adult (RPKM 7.7), cortex adult (RPKM 7.2) and 21 other tissues See more Orthologs human all
Genomic context
Location: 5; 5 C1 See Pcdh7 in Genome Data Viewer
Exon count: 7
Annotation release Status Assembly Chr Location
108 current GRCm38.p6 (GCF_000001635.26) 5 NC_000071.6 (57716145..58133236)
Build 37.2 previous assembly MGSCv37 (GCF_000001635.18) 5 NC_000071.5 (58109260..58523479)
Chromosome 5 - NC_000071.6
Page 6 of 9 https://www.alphaknockout.com
Transcript information: This gene has 7 transcripts
Gene: Pcdh7 ENSMUSG00000029108
Description protocadherin 7 [Source:MGI Symbol;Acc:MGI:1860487] Gene Synonyms BH-protocadherin Location Chromosome 5: 57,717,967-58,133,230 forward strand. GRCm38:CM000998.2 About this gene This gene has 7 transcripts (splice variants), 198 orthologues, 33 paralogues and is a member of 1 Ensembl protein family. Transcripts
Name Transcript ID bp Protein Translation ID Biotype CCDS UniProt Flags
Pcdh7- ENSMUST00000191837.5 8785 1255aa ENSMUSP00000142319.1 Protein coding CCDS51505 A0A0A6YY83 TSL:1 203 GENCODE basic APPRIS ALT1
Pcdh7- ENSMUST00000068110.9 5611 1069aa ENSMUSP00000066306.7 Protein coding CCDS19296 A2RS43 TSL:1 201 GENCODE basic APPRIS P3
Pcdh7- ENSMUST00000094783.6 3752 1247aa ENSMUSP00000092376.4 Protein coding CCDS80286 E9Q2S0 TSL:1 202 GENCODE basic APPRIS ALT1
Pcdh7- ENSMUST00000192287.5 2602 802aa ENSMUSP00000142276.1 Protein coding - A0A0A6YY48 CDS 5' 205 incomplete TSL:1
Pcdh7- ENSMUST00000192048.1 863 125aa ENSMUSP00000141505.1 Protein coding - A0A0A6YWD5 CDS 5' 204 incomplete TSL:3
Pcdh7- ENSMUST00000199310.1 659 171aa ENSMUSP00000143387.1 Protein coding - A0A0G2JG16 TSL:2 207 GENCODE basic
Pcdh7- ENSMUST00000195156.2 2199 379aa ENSMUSP00000141378.1 Nonsense mediated - A0A0A6YW34 CDS 5' 206 decay incomplete TSL:5
Page 7 of 9 https://www.alphaknockout.com
435.26 kb Forward strand 57.8Mb 57.9Mb 58.0Mb 58.1Mb Genes (Comprehensive set... Pcdh7-203 >protein coding
Pcdh7-201 >protein codinGgm42481-201 >TEC Gm17977-201 >transcribed processed pseudogene Gm42483-201 >TEC
Pcdh7-202 >protein coding
Pcdh7-205 >protein coding Gm42479-201 >TEC Gm37720-201 >TEC Gm42486-201 >TEC
Pcdh7-206 >nonsense mediated decay
Pcdh7-207 >protein coding Gm42478-201 >TEC
Pcdh7-204 >protein coding
Gm42635-201 >TEC Gm42480-201 >TEC Gm43715-201 >TEC Gm42640-201 >TEC
Gm42482-201 >TEC Gm42639-201 >TEC
Gm42484-201 >TEC
Contigs AC163275.4 > AC131036.4 > < AC119244.12 < AC125025.7
Genes < 4932441J04Rik-201lncRNA (Comprehensive set...
< 4932441J04Rik-202retained intron
Regulatory Build
57.8Mb 57.9Mb 58.0Mb 58.1Mb Reverse strand 435.26 kb
Regulation Legend
CTCF Enhancer Open Chromatin Promoter Promoter Flank
Gene Legend Protein Coding
Ensembl protein coding merged Ensembl/Havana
Non-Protein Coding
processed transcript pseudogene RNA gene
Page 8 of 9 https://www.alphaknockout.com
Transcript: ENSMUST00000191837
415.26 kb Forward strand
Pcdh7-203 >protein coding
ENSMUSP00000142... Transmembrane heli... MobiDB lite Low complexity (Seg) Cleavage site (Sign... Superfamily Cadherin-like superfamily SMART Cadherin-like Prints Cadherin-like Pfam Cadherin, N-terminal Cadherin-like Protocadherin
PROSITE profiles PS50268 PROSITE patterns Cadherin conserved site PANTHER PTHR24028:SF253
PTHR24028 Gene3D 2.60.40.60 CDD cd11304
All sequence SNPs/i... Sequence variants (dbSNP and all other sources)
Variant Legend missense variant synonymous variant
Scale bar 0 200 400 600 800 1000 1255
We wish to acknowledge the following valuable scientific information resources: Ensembl, MGI, NCBI, UCSC.
Page 9 of 9