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Mouse Med11 Conditional Knockout Project (CRISPR/Cas9)
Objective: To create a Med11 conditional knockout Mouse model (C57BL/6J) by CRISPR/Cas-mediated genome engineering.
Strategy summary: The Med11 gene (NCBI Reference Sequence: NM_025397 ; Ensembl: ENSMUSG00000018923 ) is located on Mouse chromosome 11. 3 exons are identified, with the ATG start codon in exon 1 and the TGA stop codon in exon 3 (Transcript: ENSMUST00000019067). Exon 1~2 will be selected as conditional knockout region (cKO region). Deletion of this region should result in the loss of function of the Mouse Med11 gene. To engineer the targeting vector, homologous arms and cKO region will be generated by PCR using BAC clone RP23-87B1 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 1~2 covers 61.54% of the coding region. Start codon is in exon 1, and stop codon is in exon 3. The size of intron 2 for 3'-loxP site insertion: 763 bp. The size of effective cKO region: ~620 bp. The cKO region does not have any other known gene.
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Overview of the Targeting Strategy
gRNA region
Wildtype allele A gRNA region T
5' G 3'
1 2 3 5
Targeting vector A T G
Targeted allele A T G
Constitutive KO allele (After Cre recombination)
Legends Homology arm Exon of mouse Med11 cKO region Exon of mouse Cxcl16 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. Tandem repeats are found in the dot plot matrix. It may be difficult to construct this targeting vector.
Overview of the GC Content Distribution Window size: 300 bp
Sequence 12
Summary: Full Length(6860bp) | A(28.16% 1932) | C(23.35% 1602) | T(24.55% 1684) | G(23.94% 1642)
Note: The sequence of homologous arms and cKO region is analyzed to determine the GC content. Significant high GC-content regions are found. It may be difficult to construct this targeting vector.
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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% chr11 + 70448980 70451979 3000 browser details YourSeq 419 213 1089 3000 93.3% chr14 - 20124209 20560304 436096 browser details YourSeq 240 538 1095 3000 87.2% chr5 - 109660594 109661186 593 browser details YourSeq 238 565 1082 3000 85.0% chrX - 106173651 106173998 348 browser details YourSeq 218 213 716 3000 87.1% chr16 - 31982484 31982884 401 browser details YourSeq 199 5 353 3000 86.8% chr10 + 91136407 91136716 310 browser details YourSeq 197 623 1112 3000 86.9% chr12 + 66213076 66213562 487 browser details YourSeq 166 462 1096 3000 82.0% chr9 + 72365636 72366018 383 browser details YourSeq 165 667 1096 3000 83.2% chr10 + 37061410 37061734 325 browser details YourSeq 163 901 1112 3000 91.5% chr2 - 148131803 148132032 230 browser details YourSeq 161 564 1063 3000 84.7% chr18 - 37909723 37910158 436 browser details YourSeq 158 916 1108 3000 91.8% chrX + 71505877 71506092 216 browser details YourSeq 158 668 1112 3000 80.9% chr2 + 68827084 68827341 258 browser details YourSeq 157 909 1402 3000 90.9% chr15 - 17805245 17805793 549 browser details YourSeq 156 895 1112 3000 88.5% chr7 - 114176830 114177068 239 browser details YourSeq 156 916 1111 3000 90.9% chr15 + 17126520 17126738 219 browser details YourSeq 154 916 1112 3000 91.1% chr19 + 36587521 36587738 218 browser details YourSeq 154 911 1112 3000 90.6% chr11 + 72904209 72904444 236 browser details YourSeq 152 920 1112 3000 92.4% chr1 - 187569822 187570036 215 browser details YourSeq 152 916 1112 3000 93.2% chr6 + 84081534 84081751 218
Note: The 3000 bp section upstream 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 3000 1 3000 3000 100.0% chr11 + 70452590 70455589 3000 browser details YourSeq 353 2097 2467 3000 97.6% chr11 + 70454646 70455016 371 browser details YourSeq 349 2061 2427 3000 97.6% chr11 + 70454690 70455056 367 browser details YourSeq 191 2308 2518 3000 93.7% chr11 + 70454617 70454822 206 browser details YourSeq 144 2347 2500 3000 96.8% chr11 + 70454616 70454769 154 browser details YourSeq 144 2027 2180 3000 96.8% chr11 + 70454936 70455089 154 browser details YourSeq 102 2027 2140 3000 94.8% chr11 + 70454976 70455089 114 browser details YourSeq 89 2392 2500 3000 95.9% chr11 + 70454621 70454729 109 browser details YourSeq 40 956 1022 3000 84.7% chr4 - 98558225 98558290 66 browser details YourSeq 40 1486 1557 3000 77.8% chr11 - 54697041 54697112 72 browser details YourSeq 40 1482 1610 3000 68.9% chr10 - 94932555 94932681 127 browser details YourSeq 39 1481 1545 3000 91.5% chr13 + 114618538 114618602 65 browser details YourSeq 37 1893 1973 3000 75.4% chr10 - 128063090 128063171 82 browser details YourSeq 36 1892 1939 3000 95.3% chr1 + 30956160 30956390 231 browser details YourSeq 34 2027 2060 3000 100.0% chr11 + 70455056 70455089 34 browser details YourSeq 33 955 1032 3000 84.7% chr4 + 117851396 117851471 76 browser details YourSeq 33 2468 2500 3000 100.0% chr11 + 70454617 70454649 33 browser details YourSeq 30 1480 1543 3000 73.5% chr1 - 171187366 171187429 64 browser details YourSeq 29 2000 2041 3000 96.9% chr3 + 89847061 89847103 43 browser details YourSeq 29 1909 1939 3000 90.0% chr1 + 155076639 155076668 30
Note: The 3000 bp section downstream of Exon 2 is BLAT searched against the genome. No significant similarity is found.
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Gene and protein information: Med11 mediator complex subunit 11 [ Mus musculus (house mouse) ] Gene ID: 66172, updated on 12-Aug-2019
Gene summary
Official Symbol Med11 provided by MGI Official Full Name mediator complex subunit 11 provided by MGI Primary source MGI:MGI:1913422 See related Ensembl:ENSMUSG00000018923 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 AI465144; AW545069; 1110030J09Rik Expression Ubiquitous expression in CNS E11.5 (RPKM 12.5), placenta adult (RPKM 11.9) and 28 other tissues See more Orthologs human all
Genomic context
Location: 11; 11 B3 See Med11 in Genome Data Viewer
Exon count: 3
Annotation release Status Assembly Chr Location
108 current GRCm38.p6 (GCF_000001635.26) 11 NC_000077.6 (70451918..70453732)
Build 37.2 previous assembly MGSCv37 (GCF_000001635.18) 11 NC_000077.5 (70265433..70267229)
Chromosome 11 - NC_000077.6
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Transcript information: This gene has 3 transcripts
Gene: Med11 ENSMUSG00000018923
Description mediator complex subunit 11 [Source:MGI Symbol;Acc:MGI:1913422] Gene Synonyms 1110030J09Rik Location Chromosome 11: 70,451,919-70,453,727 forward strand. GRCm38:CM001004.2 About this gene This gene has 3 transcripts (splice variants), 162 orthologues, is a member of 1 Ensembl protein family and is associated with 9 phenotypes. Transcripts
Name Transcript ID bp Protein Translation ID Biotype CCDS UniProt Flags
Med11-201 ENSMUST00000019067.7 902 117aa ENSMUSP00000019067.7 Protein coding CCDS24947 Q9D8C6 TSL:1 GENCODE basic APPRIS P1
Med11-202 ENSMUST00000151013.7 445 127aa ENSMUSP00000134323.1 Protein coding - G3UZ31 TSL:2 GENCODE basic
Med11-203 ENSMUST00000152348.1 870 No protein - Retained intron - - TSL:2
21.81 kb Forward strand 70.445Mb 70.450Mb 70.455Mb 70.460Mb Genes (Comprehensive set... Med11-201 >protein coding Zmynd15-203 >protein coding
Med11-202 >protein coding Zmynd15-201 >protein coding
Med11-203 >retained intron Zmynd15-206 >protein coding
Zmynd15-204 >protein coding
Zmynd15-205 >retained intron
Zmynd15-202 >protein coding
Contigs AL596096.7 >
Genes < Cxcl16-201protein coding (Comprehensive set...
< Cxcl16-202nonsense mediated decay
< Cxcl16-203retained intron
Regulatory Build
70.445Mb 70.450Mb 70.455Mb 70.460Mb Reverse strand 21.81 kb
Regulation Legend CTCF Promoter Promoter Flank Transcription Factor Binding Site
Gene Legend Protein Coding
Ensembl protein coding merged Ensembl/Havana
Non-Protein Coding
processed transcript
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Transcript: ENSMUST00000019067
1.81 kb Forward strand
Med11-201 >protein coding
ENSMUSP00000019... Coiled-coils (Ncoils) Pfam Mediator complex, subunit Med11 PANTHER Mediator complex, subunit Med11
PTHR22890:SF2
All sequence SNPs/i... Sequence variants (dbSNP and all other sources)
Variant Legend missense variant synonymous variant
Scale bar 0 10 20 30 40 50 60 70 80 90 100 117
We wish to acknowledge the following valuable scientific information resources: Ensembl, MGI, NCBI, UCSC.
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