Full-Length & Uncut Full-Length cDNA Library Services
Full-Length Inserts
Finding full-length target sequences is a key objective of many library screens. Our novel full-length cDNA synthesis procedure combines a proprietary process for reducing RNase H reverse transcriptase activity with unique techniques for mRNA isolation, 5´ cap full-length enrichment, and reduction of oligo(dT) priming. Together, these steps greatly enhance the likelihood of constructing a library with full-length inserts (Figure 1 and Table 1), giving you better targets at which to aim.
Uncut Libraries and the Power of Gateway® Technology
To further enhance the chances of identifying full-length inserts, we have combined our proprietary full-length library procedure with the Gateway® recombination-based cloning technology. Using Gateway® technology allows us to avoid restriction enzyme digestion of the cDNA during library construction. Stringent quality control ensures that libraries are highly enriched for full-length cDNA clones (Figure 1 and Table 1). Our full-length cDNA libraries routinely achieve:
- 65–85% full-length clones (Table 2)
- >1 x 107 clones
- Insert lengths up to 8 kb
Figure 1. Estimation of the percentage of full-length inserts in six different cDNA libraries produced using Invitrogen’s proprietary full-length cDNA library technologies.
Table 1. Sample data for full-length human brain library. N=20
| Gene | Size(kb) | Full-length clones |
|---|---|---|
| EIF4E | 2.0 | 100 |
| Protein tyrosine kinase | 3.0 | 71 |
| TOP 3 | 3.7 | 80 |
| Helix loop helix | 4.0 | 80 |
| 20XDH | 4.1 | 81 |
| MAP4 | 5.5 | 72 |
Table 2. Comparison of uncut full-length (FL) and full-length (FL) cDNA libraries.
| Major Difference | Uncut FL Library | FL Library |
|---|---|---|
| Vector | PENTR222 | PCMVSport6.1 or pcDNA3.1 |
| Starting Material (mRNA) | 2-6ug | 15ug |
| Res. Enzyme (Not I) Cut | No | Yes |
| Cloning Strategy | BxP Recombination | Ligation |
| Normalizable | No | Yes |
For more information or to order a service, call 800.955.6288 x2 or e-mail custom.services@lifetech.com.
