https://www.alphaknockout.com
Mouse Fibp Conditional Knockout Project (CRISPR/Cas9)
Objective: To create a Fibp conditional knockout Mouse model (C57BL/6J) by CRISPR/Cas-mediated genome engineering.
Strategy summary: The Fibp gene (NCBI Reference Sequence: NM_001253832 ; Ensembl: ENSMUSG00000024911 ) is located on Mouse chromosome 19. 10 exons are identified, with the ATG start codon in exon 1 and the TAA stop codon in exon 10 (Transcript: ENSMUST00000025847). Exon 4 will be selected as conditional knockout region (cKO region). Deletion of this region should result in the loss of function of the Mouse Fibp gene. To engineer the targeting vector, homologous arms and cKO region will be generated by PCR using BAC clone RP23-63O6 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:
Exon 4 starts from about 37.73% of the coding region. The knockout of Exon 4 will result in frameshift of the gene. The size of intron 3 for 5'-loxP site insertion: 1015 bp, and the size of intron 4 for 3'-loxP site insertion: 538 bp. The size of effective cKO region: ~601 bp. The cKO region does not have any other known gene.
Page 1 of 8 https://www.alphaknockout.com
Overview of the Targeting Strategy
Wildtype allele gRNA region 5' gRNA region 3'
1 2 3 4 5 6 7 8 9 10 Targeting vector
Targeted allele
Constitutive KO allele (After Cre recombination)
Legends Homology arm Exon of mouse Fibp cKO region loxP site
Page 2 of 8 https://www.alphaknockout.com
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(7101bp) | A(23.7% 1683) | C(25.98% 1845) | T(23.88% 1696) | G(26.43% 1877)
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.
Page 3 of 8 https://www.alphaknockout.com
BLAT Search Results (up)
QUERY SCORE START END QSIZE IDENTITY CHROM STRAND START END SPAN ------browser details YourSeq 3000 1 3000 3000 100.0% chr19 + 5459283 5462282 3000 browser details YourSeq 102 268 435 3000 90.0% chr11 - 64175755 64175940 186 browser details YourSeq 100 268 381 3000 94.8% chr7 + 25962599 25962718 120 browser details YourSeq 99 266 381 3000 93.2% chr3 - 144382761 144382882 122 browser details YourSeq 98 262 382 3000 91.6% chr8 + 112364858 112364983 126 browser details YourSeq 97 268 381 3000 93.0% chr2 - 29709742 29709861 120 browser details YourSeq 96 268 382 3000 93.0% chr5 - 14036252 14036373 122 browser details YourSeq 96 271 381 3000 93.7% chr18 + 75438793 75438909 117 browser details YourSeq 95 254 382 3000 88.7% chr17 - 77732260 77732388 129 browser details YourSeq 95 268 381 3000 92.2% chr13 - 37743465 37743584 120 browser details YourSeq 95 271 382 3000 93.7% chr7 + 127043581 127043699 119 browser details YourSeq 93 271 381 3000 93.5% chr7 - 27648641 27648757 117 browser details YourSeq 93 268 381 3000 91.3% chr16 - 79820307 79820426 120 browser details YourSeq 93 267 380 3000 91.2% chr12 + 39595696 39595815 120 browser details YourSeq 93 268 381 3000 88.4% chr1 + 131914776 131914887 112 browser details YourSeq 92 268 381 3000 91.1% chr14 - 86804836 86804955 120 browser details YourSeq 91 272 381 3000 91.9% chr3 - 105080637 105080752 116 browser details YourSeq 91 268 381 3000 90.4% chr17 - 90235624 90235743 120 browser details YourSeq 91 268 381 3000 90.4% chr4 + 5117479 5117598 120 browser details YourSeq 91 274 381 3000 92.6% chr19 + 5834138 5834251 114
Note: The 3000 bp section upstream of Exon 4 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% chr19 + 5462884 5465883 3000 browser details YourSeq 56 1866 1972 3000 84.0% chr12 + 91565491 91565756 266 browser details YourSeq 53 1762 2133 3000 86.5% chr16 + 5177031 5177629 599 browser details YourSeq 46 1870 1939 3000 75.5% chr16 - 56147477 56147533 57 browser details YourSeq 45 1752 1913 3000 92.4% chr11 + 109285493 109285794 302 browser details YourSeq 43 1866 1933 3000 86.7% chr1 - 160549575 160549652 78 browser details YourSeq 42 1866 1916 3000 92.2% chr11 + 101461601 101513979 52379 browser details YourSeq 41 1866 1914 3000 91.9% chr13 - 43221688 43221736 49 browser details YourSeq 41 1866 1913 3000 93.8% chr15 + 64063190 64063240 51 browser details YourSeq 37 1866 1912 3000 89.4% chr2 - 91295854 91295900 47 browser details YourSeq 37 1866 1931 3000 69.7% chr11 - 3158737 3158792 56 browser details YourSeq 37 1867 1913 3000 89.4% chr10 + 117207766 117207812 47 browser details YourSeq 36 1866 1925 3000 84.7% chr11 - 20280379 20280442 64 browser details YourSeq 36 1867 1912 3000 84.5% chr8 + 60826896 60826940 45 browser details YourSeq 35 1892 1998 3000 89.8% chr12 - 101009670 101009775 106 browser details YourSeq 35 1866 1913 3000 87.5% chr11 - 114744121 114744171 51 browser details YourSeq 35 1866 1914 3000 85.8% chr13 + 29364634 29364682 49 browser details YourSeq 34 1866 1909 3000 88.7% chr11 - 98006784 98006827 44 browser details YourSeq 34 1866 1913 3000 85.5% chr10 - 7665919 7665966 48 browser details YourSeq 34 1866 1909 3000 88.7% chr2 + 157365906 157365949 44
Note: The 3000 bp section downstream of Exon 4 is BLAT searched against the genome. No significant similarity is found.
Page 4 of 8 https://www.alphaknockout.com
Gene and protein information: Fibp fibroblast growth factor (acidic) intracellular binding protein [ Mus musculus (house mouse) ] Gene ID: 58249, updated on 24-Oct-2019
Gene summary
Official Symbol Fibp provided by MGI Official Full Name fibroblast growth factor (acidic) intracellular binding protein provided by MGI Primary source MGI:MGI:1926233 See related Ensembl:ENSMUSG00000024911 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 2010004G08Rik; 2010005N21Rik; 3010027N18Rik Expression Ubiquitous expression in large intestine adult (RPKM 48.9), CNS E14 (RPKM 48.4) and 28 other tissues See more Orthologs human all
Genomic context
Location: 19; 19 A See Fibp in Genome Data Viewer
Exon count: 10
Annotation release Status Assembly Chr Location
108 current GRCm38.p6 (GCF_000001635.26) 19 NC_000085.6 (5460598..5465052)
Build 37.2 previous assembly MGSCv37 (GCF_000001635.18) 19 NC_000085.5 (5460694..5464994)
Chromosome 19 - NC_000085.6
Page 5 of 8 https://www.alphaknockout.com
Transcript information: This gene has 13 transcripts
Gene: Fibp ENSMUSG00000024911
Description fibroblast growth factor (acidic) intracellular binding protein [Source:MGI Symbol;Acc:MGI:1926233] Gene Synonyms 2010004G08Rik, 2010005N21Rik, 3010027N18Rik Location Chromosome 19: 5,460,615-5,465,051 forward strand. GRCm38:CM001012.2 About this gene This gene has 13 transcripts (splice variants), 181 orthologues and is a member of 1 Ensembl protein family. Transcripts
Name Transcript ID bp Protein Translation ID Biotype CCDS UniProt Flags
Fibp-201 ENSMUST00000025847.6 1215 364aa ENSMUSP00000025847.5 Protein coding CCDS57130 Q8K2D8 TSL:1 GENCODE basic
Fibp-205 ENSMUST00000225141.1 1209 357aa ENSMUSP00000153409.1 Protein coding - Q9JI19 GENCODE basic APPRIS P1
Fibp-207 ENSMUST00000225796.1 2277 No protein - Retained intron - - -
Fibp-208 ENSMUST00000225917.1 1923 No protein - Retained intron - - -
Fibp-211 ENSMUST00000237181.1 1569 No protein - Retained intron - - -
Fibp-206 ENSMUST00000225222.1 1499 No protein - Retained intron - - -
Fibp-212 ENSMUST00000237258.1 1049 No protein - Retained intron - - -
Fibp-204 ENSMUST00000224944.1 773 No protein - Retained intron - - -
Fibp-213 ENSMUST00000238011.1 730 No protein - Retained intron - - -
Fibp-209 ENSMUST00000226017.1 721 No protein - Retained intron - - -
Fibp-203 ENSMUST00000224422.1 600 No protein - Retained intron - - -
Fibp-202 ENSMUST00000224005.1 410 No protein - Retained intron - - -
Fibp-210 ENSMUST00000235533.1 278 No protein - Retained intron - - -
Page 6 of 8 https://www.alphaknockout.com
24.44 kb Forward strand 5.455Mb 5.460Mb 5.465Mb 5.470Mb 5.475Mb Genes (Comprehensive set... Fosl1-201 >protein coding Fibp-211 >retained intron Efemp2-207 >retained intron
Fibp-205 >protein coding Efemp2-216 >retained intron
Fibp-208 >retained intron Efemp2-212 >retained intron
Fibp-207 >retained intron Fibp-210 >retained intron Efemp2-206 >protein coding
Fibp-201 >protein coding Efemp2-208 >protein coding
Fibp-204 >retained intron Efemp2-215 >protein coding
Fibp-213 >retained intron Efemp2-204 >protein coding
Fibp-206 >retained intron Efemp2-209 >protein coding
Fibp-212 >retained intron Efemp2-205 >protein coding
Fibp-203 >retained intron Efemp2-211 >protein coding
Fibp-202 >retained intron Efemp2-201 >protein coding
Fibp-209 >retained intron Efemp2-217 >protein coding
Contigs AC122861.4 > < AC126029.3 Genes < Ccdc85b-201protein coding < Ctsw-202retained intron (Comprehensive set...
< Ccdc85b-202protein coding < Ctsw-201protein coding
< Ctsw-203retained intron
< Ctsw-205protein coding
< Ctsw-206retained intron
< Ctsw-204retained intron
Regulatory Build
5.455Mb 5.460Mb 5.465Mb 5.470Mb 5.475Mb Reverse strand 24.44 kb
Regulation Legend CTCF Open Chromatin Promoter Promoter Flank
Gene Legend Protein Coding
merged Ensembl/Havana Ensembl protein coding
Non-Protein Coding
processed transcript
Page 7 of 8 https://www.alphaknockout.com
Transcript: ENSMUST00000025847
4.42 kb Forward strand
Fibp-201 >protein coding
ENSMUSP00000025... Low complexity (Seg) Pfam Acidic fibroblast growth factor intracellular-binding protein PANTHER Acidic fibroblast growth factor intracellular-binding protein
All sequence SNPs/i... Sequence variants (dbSNP and all other sources)
Variant Legend
missense variant synonymous variant
Scale bar 0 40 80 120 160 200 240 280 320 364
We wish to acknowledge the following valuable scientific information resources: Ensembl, MGI, NCBI, UCSC.
Page 8 of 8