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Invitrogen™ Retic Lysate IVT™ Kit
Description
Includes
Retic Lysate should be stored at -80°C. 20X Translation Mix minus-Met, 20X Translation Mix minus-Leu, High Salt Mix minus-Met, Low Salt Mix minus-Met, and Capped Xef-1 RNA should be stored below -70°C. Nuclease-free Water can be stored at any temperature.
The Retic Lysate IVT™ Kit efficiently translates in vitro-synthesized transcripts, poly(A) RNA, and total RNA, including difficult-to-translate RNAs. The optimized lysate results in high protein yield, biological activity, and consistent performance. A report in Nucleic Acids Research (1) found that some commercial reticulocyte lysates had poor fidelity for initiation at the appropriate AUG initiator codon. The Retic Lysate IVT Kit was tested using Kozak's constructs and it demonstrated high fidelity for initiating at the proper AUG.
- Efficiently translates a wide variety of high molecular weight proteins and both capped and uncapped mRNAs
- Accommodates large volume of template mRNA
The lysate has been optimized for translation by the addition of an ATP-regenerating system, hemin, and calf liver tRNA. Included in the kit is an optimized Translation Mix (-Met or -Leu) for the translation of both capped and uncapped messages along with High and Low Salt buffers. These buffers can be used to maximize protein yield and activity from difficult-to-translate sequences.
1. Kozak, M. (1990) Evaluation of the fidelity of initiation of translation in reticulocyte lysates from commercial sources. Nucleic Acids Res 18:2828.
Specifications
Specifications
| Content And Storage | • 5 × 200 μl (store at -80°C) • 2 × 1.75 ml Nuclease-free Water (store at any temperature) • 100 μl 20X Translation Mix minus-Met • 100 μl 20X Translation Mix minus-Leu • 100 μl High Salt Mix minus-Met • 100 μl Low Salt Mix minus-Met • 50 μl Capped Xef-1 RNA (50 μg/ml) Unless otherwise stated, store reagents below -70°C. Do not store cell-free translation systems in the liquid phase of a liquid nitrogen storage system. |
| Format | Kit |
| Expressed Protein Type | Water Soluble Protein |
| Expression Mechanism | Cell-Free Expression |
| Expression System | Cell-Free |
| For Use With (Application) | In Vitro Transcription |
| No. of Reactions | 60 Reactions |
| Product Line | Ambion |
| Product Type | Retic Lysate |
| Quantity | 60 Reactions |
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Frequently Asked Questions (FAQs)
Rabbit Reticulocyte Lysate (RRL) and Wheat Germ Lysate (WGE) can often be used interchangeably for in vitro translation. In general, RRL efficiently translates proteins greater than 30 kDa while WGE works well for proteins of all sizes (including those in the range of 15-30 kDa), although larger proteins may be expressed more efficiently in RRL. This appears to be partially due to the relative instability of small proteins in RRL. However, certain templates are translated more efficiently in RRL, whereas others produce more protein in WGE; the difference in yield can be quite dramatic and must be determined empirically.
Another consideration is whether the protein of interest is present endogenously in either RRL or WGE, or whether factors that might interfere with downstream applications may be present in RRL or WGE. This can be determined by performing a control in vitro translation reaction with no template added. The RRL is recommended if Canine Pancreatic Microsomal Membranes are used for cotranslational processing (core glycosylation or signal peptide cleavage) or for production of proteins that require membrane insertion for in vitro translation.
For Research Use Only. Not for use in diagnostic procedures.