Down-regulating gene expression is fundamentally important to the study of gene function. In higher eukaryotes, which have been historically difficult to manipulate for making complete genetic knock outs, researchers have used RNA interference (RNAi), which reduces gene expression post-transcriptionally without making permanent changes in the chromosome, irrespective of the chromatin structure or DNA methylation state of the target locus. RNAi causes “knock down”, reducing but not completely eliminating gene expression. Knock down can be advantageous over knock out in applications where complete ablation of gene expression is lethal, or in cases where one wants downregulation to be conditional. In contrast to short interfering RNAs (siRNAs), which knock genes down transiently, short hairpin RNAs (shRNAs) can be integrated into the chromosome to downregulate gene expression stably or conditionally.

GeneCopoeia offers the generation of cell lines stably expressing shRNAs targeting a gene of interest. We will design your shRNAs and integrate them into virtually any cell line you choose.

• Determination of gene function in cells
• Expression profiling to identify candidate downstream genes in a network or pathway
• Drug target validation

• Lentifect™ purified lentiviral particles
• OmicsLink™ shRNA clones
• Premade labeled cancer cell lines

Genome editing-the ability to make specific changes at targeted genomic sites in complex organisms-is of fundamental importance in biology and medicine. Over the past several years, researchers have developed chimeric DNA binding proteins complexed with nucleases to stimulate double-strand breaks at defined genomic loci, allowing the ability to insert, delete, and replace genetic information at will. Recently, the Clustered, Regularly Interspersed, Short Palindromic Repeats (CRISPR) systems have gained favor as genome editing tools.

Along with CRISPR-based plasmid design and construction services, GeneCopoeia offers the generation of stable cell lines carrying specific, targeted modifications created with CRISPR. In addition, genes can be stably activated or repressed using CRISPR activation (CRISPRa) or CRISPR inhibition (CRISPRi), respectively. We will design the CRISPR target site, and, if needed, donor constructs used for homologous recombination-mediated applications, transfect your cell line of choice, and isolate single or double allele-modified clones. Our stable cell line service can also be used with our human and mouse Safe Harbor Integration system and knock-in ORF clones.

• Gene knock out
• Introduction of point mutations or defined insertions/deletions
• Correction of disease mutations back to wild type
• Transgene knock in at a customer-specified locus or at human and mouse Safe Harbor sites
• Gene tagging
• Promoter or gene replacement
• Gene activation
• Gene repression

• GeneHero™ CRISPR products and services
• Human AAVS1 and mouse Rosa26 safe harbor gene targeting kits and clones
• CRISPR-Cas9 stable cell lines
• CRISPRa stable cell lines
• CRISPRi stable cell lines
• Premade labeled cancer cell lines
• OncoSpot™ cancer biomarker mutant cell lines

Primary cells undergo a finite number of population doublings and reach a state called replicative senescence after a certain number of divisions that limits the proliferation of aged or damaged cells. In order to make primary cell lines continue to grow and divide indefinitely in vitro, an immortalization process is applied to primary cells to significantly prolong their lifespan. Immortalized cells are important models for studies in cell biology, cellular metabolism and differentiation; and can be used for research in cancer biology, immunology, neurology, hematology and more.

SV40T – The most reliable and simplest way to immortalize many types of primary cells. The viral gene SV40 large T antigen is introduced into primary cells to override the cell cycle by inactivating tumor suppressor genes that induce cellular senescence.

hTERT – One of the most popular approaches to cell immortalization. The telomerase reverse transcriptase (TERT) protein is inactive in most somatic cells, causing the length of telomeres to shorten with age, leading to senescence. Expression of hTERT prevents the telomere from being truncated during cell division, thus cells are able to avoid replicative senescence. 

Fluorescence in situ hybridization (FISH)

For chromosome enumeration, as well as detection of disease-related genetic mutations, amplification and rearrangements resulting in large-scale changes in the physical arrangement of genes on chromosomes. More details…

Chromosome enumeration FISH probes can be used to determine chromosome copy number in diagnosis and prognosis of cancers and other diseases.

Disease gene FISH probes can be used to detect oncogene deletion, amplification and rearrangement in cancer cellls.

STR analysis

Cross-contamination among cultured cell lines is a prevalent and persistent  problem. Short Tandom Repeat (STR) DNA profiling identifies polymorphisms among STR markers; it is a relatively easy, low cost and reliable method for the detection of cellular cross-contamination to reveal true cell line identity.

MSI analysis

MSI is failure to correct the errors that spontaneously occur during DNA replication, resulting in the accumulation of chromosomal mutations at certain repetitive DNA motifs as microsatellites. MSI testing is performed to analyze colon and other tumor tissue samples for features suggestive of Lynch syndrome or hereditary non-polyposis colorectal cancer (HNPCC).

RT-qPCR

For the quantification of alternative splicing variants and gene expression profiling. Custom qPCR primers designed for the quantification of mRNA and detection of exon skipping, intron retention, alternative 5' or 3' splicing and other splicing variants to identify and quantify different splicing variants.

Mycoplasma detection

Mycoplasma contamination of cell cultures can easily go undetected and adversely affect cell physiology and metabolism. Therefore, routine testing is recommended to ensure the purity of cultured cells. GeneCopoeia offers Mycoplasma Detection Services as a preliminary testing step prior to all mammalian stable cell line services using our own highly sensitive mycoplasma detection kit.