Dh5α Competent Cells, supplied by New England Biolabs, used in various techniques. Bioz Stars score: 98/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/dh5α competent cells/product/New England Biolabs
Average 98 stars, based on 1 article reviews
Price from $9.99 to $1999.99
1) Product Images from "Novel Loss-of-Function Variants in CDC14A are Associated with Recessive Sensorineural Hearing Loss in Iranian and Pakistani Patients"
Article Title: Novel Loss-of-Function Variants in CDC14A are Associated with Recessive Sensorineural Hearing Loss in Iranian and Pakistani Patients
Journal: International Journal of Molecular Sciences
Figure Legend Snippet: Characterization of the CDC14A c.1421+2T > C exchange via a minigene assay. ( a ) A schematic of the pSPL3 exon trapping vector with cloned CDC14A exon 14 (blue) and flanking sequence containing Xho I and Bam HI restriction sites that was directly amplified from proband and wild type genomic DNA. Exons A and B (purple) originate from the vector. A schematic of the resulting splice products is shown, with the wild type splicing profile (top) and splice variant sequence that activates a cryptic splice site (bottom, red asterisk). The PCR primers that were used to amplify the Exon A splice donor region (SD6) and Exon B splice acceptor region (SA2) are depicted by green arrows. ( b ) Electrophoretic visualization of cDNA RT-PCR products amplified from constructs after transfection into HEK293T cells. Amplicons were resolved on a 1% agarose gel. Wild type splicing (lane: pSPL3 CDC14A WT) yields a 380 bp product that constitutes the Exon A, exon 14 and Exon B amplified regions. The homozygous mutant amplicon (lane: pSPL3 CDC14A hom) shows a band around 380 bp that, when sequenced, indicates a cryptic splice site activation. The empty vector shows the expected 257 bp product. ( c ) Sequencing electropherograms of the exon 14 5′ splice site boundaries for the RT-PCR products for wild type (top), mutant showing cryptic splice site activation (middle) and empty vector (bottom). ( d ) In silico splice prediction tools for the c.1421+2T > C exchange that is marked with red lines visualized with Alamut visual (2.10). The upper panel shows the reference sequence splice scores and the lower panel shows the splice scores for the c.1421+2T > C exchange with multiple in silico prediction tools estimating the loss of the native exon 14 5′ donor splice site that is due to the variant (shown with a black box). In the mutant panel, the splice scores of an adjacent cryptic 5′ donor site are either unchanged or strengthened and marked with a black arrow. ( e ) Effect of the splice variant on the protein, comparing wild type (top) and the truncated (bottom) protein resulting from the aberrantly spliced product (visualized with Mutalyzer). The amino acid residues marked in red are those that are altered due to the variant.
Techniques Used: Mini Gene Assay, Plasmid Preparation, Clone Assay, Sequencing, Amplification, Variant Assay, Polymerase Chain Reaction, Reverse Transcription Polymerase Chain Reaction, Construct, Transfection, Agarose Gel Electrophoresis, Mutagenesis, Activation Assay, In Silico