内容及储存
SuperScript® III Reverse Transcriptase is supplied with a vial of 5X first-strand buffer [250 mM Tris-HCl (pH 8.3), 375 mM KCl, 15 mM MgCl2], and a vial of 100 mM DTT.
Store at -20°C. Guaranteed stable for 6 months when properly stored.
描述SuperScript® III Reverse Transcriptase (RT) is a proprietary mutant of SuperScript® II RT that is active at 50°C and has a half-life of 220 minutes, providing increased specificity with Gene-Specific Primers (GSPs) and the highest cDNA yield of all RTs. It is ideal for RT-PCR of a specific gene or generating cDNA from total or poly (A)+ RNA sample. Like SuperScript® II, it synthesizes a complementary DNA strand from single-stranded RNA, DNA, or an RNA:DNA hybrid. SuperScript® III RT is genetically engineered by the introduction of point mutations that increase half-life, reduce RNase activity, and increase thermal stability.
• Thermostability – Half life of 220 minutes at 50°C
• Yield – Reduced RNase H activity for more full-length cDNA
• Specificity – Full activity at 50°C for increased specificity with Gene Specific Primers
• Applications – Array labeling, cDNA libraries, RT-PCR, primer extension, and 3´ and 5´ RACE
Source: Purified from E. coli expressing the pol gene of M-MLV (1, 2), mutagenized to increase thermal stability and half life and reduce RNase H activity.
Performance and Quality Testing: SDS-PAGE purity; endodeoxyribonuclease, exodeoxyribonuclease, and ribonuclease assays (Figure 2) and yield and length of cDNA product.
Unit Definition: One unit of SuperScript® III RT is the amount of enzyme required to incorporate 1 nmole of deoxyribonucleotide into acid-precipitable material in 10 min. at 37°C using poly(A)•oligo(dT)12-18 as a template•primer (3).
Unit Reaction Conditions: 50 mM Tris-HCl (pH 8.3), 40 mM KCl, 6 mM MgCl2, 1 mM DTT, 0.5 mM [3H]dTTP, 0.1 mM poly(A), 0.1 mM oligo(dT)12-18, 0.1 mg/ml BSA, and enzyme in 50 µl for 10 min. at 37°C.
