https://www.alphaknockout.com

Mouse C1qtnf3 Knockout Project (CRISPR/Cas9)

Objective: To create a C1qtnf3 knockout Mouse model (C57BL/6J) by CRISPR/Cas-mediated genome engineering.

Strategy summary: The C1qtnf3 (NCBI Reference Sequence: NM_001204134 ; Ensembl: ENSMUSG00000058914 ) is located on Mouse 15. 6 exons are identified, with the ATG start codon in exon 1 and the TGA stop codon in exon 6 (Transcript: ENSMUST00000110523). 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: Mice homozygous for a knock-out allele exhibit increased susceptibility to collagen-induced arthritis.

Exon 2 starts from about 31.77% of the coding region. Exon 2 covers 11.7% 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 6

Legends Exon of mouse C1qtnf3 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. 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 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(26.85% 537) | C(21.9% 438) | T(30.0% 600) | G(21.25% 425)

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(27.2% 544) | C(22.2% 444) | T(29.95% 599) | G(20.65% 413)

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 + 10955958 10957957 2000 browser details YourSeq 32 35 73 2000 92.4% chr1 + 36637404 36637446 43 browser details YourSeq 25 36 60 2000 100.0% chr4 + 58108064 58108088 25 browser details YourSeq 22 35 56 2000 100.0% chr17 - 93349370 93349391 22 browser details YourSeq 21 43 63 2000 100.0% chr1 - 60089697 60089717 21

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 + 10958070 10960069 2000 browser details YourSeq 109 1016 1252 2000 85.8% chr19 - 28769749 28769986 238 browser details YourSeq 106 1023 1258 2000 86.4% chr7 + 135223455 135223691 237 browser details YourSeq 105 1027 1252 2000 78.9% chr14 - 76691560 76691777 218 browser details YourSeq 101 1029 1256 2000 78.6% chr4 - 149887937 149888117 181 browser details YourSeq 99 793 1251 2000 84.2% chr9 - 98635947 98636427 481 browser details You