Effect of Ttc22 exon2-3 deficiency on gene expression in colon tissues of C57BL mouse

To inspect the funtion of Ttc22 gene in colon of Ttc22 knockout mice, high-throughput mRNA sequencing of colon from Ttc22+/+/Ttc22+/-/Ttc22-/- mice (male, 58 weeks old) was carried out. RNA-sequencing unveils that the differentially expressed genes in colon tissues from Ttc22-/- mice were mainly enriched within pathogen infection- and immuno-related system. Overall design: Total RNA was isolated from the colon tissues of Ttc22+/+/Ttc22+/-/Ttc22-/- mice (n=3) using an RNA Extraction Kit. RNA high throughput sequencing was performed by Cloud-Seq Biotech (Shanghai, China). Briefly, total RNA was used for removing the rRNAs with NEBNext rRNA Depletion Kit (New England Biolabs, Inc., Massachusetts, USA) following the manufacturer's instructions. RNA libraries were constructed by using NEBNext® Ultra™ II Directional RNA Library Prep Kit (New England Biolabs, Inc., Massachusetts,USA) according to the manufacturer's instructions. Libraries were controlled for quality and quantified using the BioAnalyzer 2100 system (Agilent Technologies, Inc., USA). Library sequencing was performed on an illumina NovaSeq 6000 instrument with 150bp paired end reads. Paired-end reads were harvested from Illumina NovaSeq 6000 sequencer, and were quality controlled by Q30. After 3' adaptor-trimming and low quality reads removing by cutadapt software (v1.9.3), the high quality clean reads were aligned to the reference genome with hisat2 software (v2.0.4). Then, guided by the Ensembl gtf gene annotation file, cuffdiff software (part of cufflinks) was used to get the gene level FPKM as the expression profiles of mRNA.