https://www.alphaknockout.com
Mouse Laptm4b Knockout Project (CRISPR/Cas9)
Objective: To create a Laptm4b knockout Mouse model (C57BL/6J) by CRISPR/Cas-mediated genome engineering.
Strategy summary: The Laptm4b gene (NCBI Reference Sequence: NM_033521 ; Ensembl: ENSMUSG00000022257 ) is located on Mouse chromosome 15. 7 exons are identified, with the ATG start codon in exon 1 and the TGA stop codon in exon 7 (Transcript: ENSMUST00000022867). Exon 2 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 2 starts from about 14.68% of the coding region. Exon 2 covers 16.45% of the coding region. The size of effective KO region: ~112 bp. The KO region does not have any other known gene.
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Overview of the Targeting Strategy
Wildtype allele 5' gRNA region gRNA region 3'
1 2 7
Legends Exon of mouse Laptm4b Knockout region
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Overview of the Dot Plot (up) Window size: 15 bp
Forward Reverse Complement
Sequence 12
Note: The 2000 bp section upstream of Exon 2 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.
Overview of the Dot Plot (down) Window size: 15 bp
Forward Reverse Complement
Sequence 12
Note: The 2000 bp section downstream of Exon 2 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.
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Overview of the GC Content Distribution (up) Window size: 300 bp
Sequence 12
Summary: Full Length(2000bp) | A(24.1% 482) | C(22.35% 447) | T(30.8% 616) | G(22.75% 455)
Note: The 2000 bp section upstream of Exon 2 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(27.05% 541) | C(17.6% 352) | T(29.5% 590) | G(25.85% 517)
Note: The 2000 bp section downstream of Exon 2 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 2000 1 2000 2000 100.0% chr15 + 34256682 34258681 2000 browser details YourSeq 77 59 196 2000 88.7% chr5 - 139124631 139124880 250 browser details YourSeq 70 139 448 2000 88.9% chr17 - 65020381 65020698 318 browser details YourSeq 53 138 208 2000 92.2% chr18 - 60659641 60659761 121 browser details YourSeq 51 137 206 2000 89.3% chr18 - 64615145 64615264 120 browser details YourSeq 44 139 206 2000 88.0% chr15 - 47079569 47079686 118 browser details YourSeq 42 139 191 2000 90.4% chr3 - 95249454 95249554 101 browser details YourSeq 41 508 735 2000 93.9% chr10 + 128367421 128367722 302 browser details YourSeq 39 166 209 2000 95.5% chr5 - 136986409 136986453 45 browser details YourSeq 35 84 122 2000 97.5% chr2 - 157402190 157402380 191 browser details YourSeq 35 655 739 2000 97.3% chr13 - 43148455 43148539 85 browser details YourSeq 34 178 223 2000 94.8% chr1 - 179288870 179288921 52 browser details YourSeq 33 708 746 2000 92.4% chr10 - 37263463 37263501 39 browser details YourSeq 32 137 195 2000 88.1% chr12 + 56167430 56167537 108 browser details YourSeq 30 709 739 2000 100.0% chr2 - 91380361 91380490 130 browser details YourSeq 30 704 735 2000 96.9% chr18 - 57405743 57405774 32 browser details YourSeq 29 709 746 2000 89.2% chr8 - 68776606 68776646 41 browser details YourSeq 28 709 738 2000 96.7% chr17 - 75145217 75145246 30 browser details YourSeq 28 180 207 2000 100.0% chr12 - 83599861 83599888 28 browser details YourSeq 27 701 731 2000 93.6% chr7 - 143105151 143105181 31
Note: The 2000 bp section upstream of Exon 2 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% chr15 + 34258794 34260793 2000 browser details YourSeq 61 1023 1163 2000 81.1% chr12 + 91522625 91522758 134 browser details YourSeq 57 1098 1231 2000 71.8% chr17 + 7537365 7537501 137 browser details YourSeq 55 1114 1229 2000 85.8% chr11 - 69485371 69498231 12861 browser details YourSeq 54 1100 1179 2000 84.7% chrX + 166781104 166781186 83 browser details YourSeq 54 1100 1182 2000 83.2% chr9 + 48278642 48278727 86 browser details YourSeq 52 1108 1185 2000 83.4% chr18 - 53651664 53651741 78 browser details YourSeq 52 1108 1179 2000 86.2% chr11 - 78296636 78296707 72 browser details YourSeq 51 1109 1179 2000 86.2% chr11 - 69275506 69275575 70 browser details YourSeq 50 1094 1354 2000 89.1% chr3 - 93495853 93496253 401 browser details YourSeq 50 1100 1184 2000 80.0% chr11 - 113605540 113605627 88 browser details YourSeq 50 1110 1193 2000 79.8% chr1 + 86186287 86186370 84 browser details YourSeq 48 1028 1155 2000 90.0% chr12 - 117270596 117270809 214 browser details YourSeq 48 1101 1178 2000 80.8% chr1 - 136272681 136272758 78 browser details YourSeq 48 1098 1179 2000 79.3% chr1 - 51858275 51858356 82 browser details YourSeq 46 1110 1214 2000 87.1% chr3 + 126984816 126984918 103 browser details YourSeq 45 1082 1150 2000 88.2% chr12 - 3253735 3253811 77 browser details YourSeq 44 1100 1214 2000 83.4% chr15 - 34302215 34302335 121 browser details YourSeq 44 1100 1179 2000 77.5% chr7 + 34136863 34136942 80 browser details YourSeq 44 1107 1206 2000 66.3% chr7 + 31231870 31231956 87
Note: The 2000 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: Laptm4b lysosomal-associated protein transmembrane 4B [ Mus musculus (house mouse) ] Gene ID: 114128, updated on 12-Aug-2019
Gene summary
Official Symbol Laptm4b provided by MGI Official Full Name lysosomal-associated protein transmembrane 4B provided by MGI Primary source MGI:MGI:1890494 See related Ensembl:ENSMUSG00000022257 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 LAPTM4beta; C330023P13Rik Expression Ubiquitous expression in adrenal adult (RPKM 150.7), ovary adult (RPKM 83.2) and 24 other tissues See more Orthologs human all
Genomic context
Location: 15; 15 B3.1 See Laptm4b in Genome Data Viewer Exon count: 7
Annotation release Status Assembly Chr Location
108 current GRCm38.p6 (GCF_000001635.26) 15 NC_000081.6 (34238026..34284295)
Build 37.2 previous assembly MGSCv37 (GCF_000001635.18) 15 NC_000081.5 (34167781..34214050)
Chromosome 15 - NC_000081.6
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Transcript information: This gene has 4 transcripts
Gene: Laptm4b ENSMUSG00000022257
Description lysosomal-associated protein transmembrane 4B [Source:MGI Symbol;Acc:MGI:1890494] Gene Synonyms C330023P13Rik Location Chromosome 15: 34,238,028-34,284,302 forward strand. GRCm38:CM001008.2 About this gene This gene has 4 transcripts (splice variants), 297 orthologues, 2 paralogues, is a member of 1 Ensembl protein family and is associated with 18 phenotypes. Transcripts
Name Transcript ID bp Protein Translation ID Biotype CCDS UniProt Flags
Laptm4b- ENSMUST00000022867.4 1902 227aa ENSMUSP00000022867.3 Protein coding CCDS27416 B2CZK6 TSL:1 201 Q91XQ6 GENCODE basic APPRIS P1
Laptm4b- ENSMUST00000226627.1 634 159aa ENSMUSP00000153935.1 Protein coding - A0A2I3BPV6 CDS 3' 203 incomplete
Laptm4b- ENSMUST00000228547.1 474 89aa ENSMUSP00000154118.1 Protein coding - A0A2I3BQL3 CDS 5' 204 incomplete
Laptm4b- ENSMUST00000226437.1 1662 44aa ENSMUSP00000154241.1 Nonsense mediated - A0A2I3BQX5 - 202 decay
66.28 kb Forward strand
Genes (Comprehensive set... Laptm4b-201 >protein coding
Laptm4b-202 >nonsense mediated decay
Laptm4b-203 >protein coding
Laptm4b-204 >protein coding
Contigs < AC133101.4 Genes < Gm18949-201processed pseudogene < Gm25809-201snoRNA (Comprehensive set...
Regulatory Build
Reverse strand 66.28 kb
Regulation Legend CTCF Enhancer Open Chromatin Promoter Promoter Flank
Gene Legend Protein Coding
Ensembl protein coding merged Ensembl/Havana
Non-Protein Coding
RNA gene processed transcript pseudogene
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Transcript: ENSMUST00000022867
46.27 kb Forward strand
Laptm4b-201 >protein coding
ENSMUSP00000022... Transmembrane heli... Low complexity (Seg) Coiled-coils (Ncoils) Pfam Lysosomal-associated transmembrane protein 4/5
PANTHER PTHR12479
Lysosomal-associated transmembrane protein 4B
All sequence SNPs/i... Sequence variants (dbSNP and all other sources)
Variant Legend
missense variant synonymous variant
Scale bar 0 20 40 60 80 100 120 140 160 180 200 227
We wish to acknowledge the following valuable scientific information resources: Ensembl, MGI, NCBI, UCSC.
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