3UTR Clone FAQs
1. Why should I use OriGene’s 3’ UTR reporter clones in the pMirTarget vector?
OriGene has the largest collection of human full-length cDNA clones. Thus OriGene can provide you 3’UTR clones rapidly and economically. Our research has shown that the pMirTarget was more sensitive when compared to other 3’UTR-luciferase reporter systems
2. Does the IRES design affect the interaction of microRNA and its targets?
No.Based on published data by Philip A. Sharp’s group at MIT and our own data, the microRNA represses the translation even if the translation is started from IRES
3. How can I get consistent, reliable results doing transient transfection?
The key is to achieve the same transfection efficiency, which requires the same amount and same quality of experimental and control DNA. In the pMirTarget, RFP expression driven by the CMV promoter is a good indicator of transfection. If your transfections (experiment and control) showed the same level of RFP signal, your 3’UTR-luciferase reporter assays will be reliable
4. What is the best positive control in a 3’ UTR reporter assays?
To our experience, the microRNA sponge or tandem repeats (5X) of mature microRNA reverse complementary sequence downstream of the luciferase stop codon in the pMirTarget serve as excellent positive controls
5. Do I have to mutate the seeding sequence to show the specificity of microRNA and its targets interaction?
Yes. It is the best way to show that the interactions are specific. OriGene offers the service for mutant clone construction. On each wildtype 3’UTR clone page, there is a clickable button of “Mutant clone request”. To place an order for a mutant clone, lick on the button and fill out the mutant sequence information. When ordered with the wildtype 3’UTR clones, the fee for the mutant clone is usually $400
6. What does your disclaimer mean?
OriGene’s disclaimer for the 3’ UTR reporter clones reads as follows: “Our molecular clone sequence data has been matched to the accession number below as a point of reference. Note that the complete sequence of our molecular clones may differ from the sequence published for this corresponding accession number, e.g., by representing a single nucleotide polymorphism (SNP).” The NCBI RefSeq mRNA sequences are continuously being revised. These sequences are therefore used only as a “reference” and not as a “standard”. OriGene’s clones are isolated from full-length cDNA libraries and may differ from the reference sequence for this reason
7. What is the 3’UTR clone Guarantee?
OriGene warrants that the product will meet the specifications listed. At OriGene’s discretion, free replacement of any non-conforming product will be made if OriGene is notified within 30 days of product receipt. If you experience any difficulty with any OriGene product, please contact our Technical Support Staff at 888-267-4436, or 301-340-3188 outside the US