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Mmachine t7 transcription kit

Manufactured by Thermo Fisher Scientific
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The MMACHINE T7 transcription kit is a laboratory equipment product designed for in vitro transcription of RNA from DNA templates. The kit provides the necessary components, including T7 RNA polymerase, ribonucleotides, and reaction buffers, to facilitate the efficient synthesis of RNA molecules.

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T7 polymerase, by Thermo Fisher Scientific (1 mentions)

Close spelling variants of this product

MMessage mMachine SP6 & T7 Transcription Kits T7 mMachine kit T7 Transcription Kit T7 kits Transcription Kits

3 protocols using mmachine t7 transcription kit

1

Reverse Genetics and Recombinant Protein Production

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BTV-1 (GenBank accession numbers FJ969719 to FJ969728) and RV SA11 (GenBank accession numbers LC333802 to LC333812) were used for RCAP and vPAR-CL studies. All mutations for reverse genetics and recombinant protein production were generated by site-directed mutagenesis. The sequences of the primers are listed in Supplementary Table S1. Transcripts for reverse genetic analyses were prepared using the mMACHINE T7 transcription kit (Thermo), following the manufacturers’ instructions.
Sung P.Y., Zhou Y., Kao C.C., Aburigh A.A., Routh A, & Roy P. (2023). A multidisciplinary approach to the identification of the protein–RNA connectome in double-stranded RNA virus capsids. Nucleic Acids Research, 51(10), 5210-5227.
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2

Recovering Mutant Bluetongue Viruses

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Site-directed mutagenesis was performed to introduce mutations into the exact copy of BTV-1 S5(VP5) or S7 (VP7) (pUC19BTV1T7S5 or pUC19BTV1T7S7) complementary DNA plasmid for virus recovery assay, as previously described14 (link),20 (link),29 (link). Briefly, BTV RNA transcripts were generated from the digested cDNA plasmid clones using a mMACHINE T7 transcription kit (Thermo Fisher). A monolayer of BSR cell (BHK-21 subclone) in six-well plates at 100% confluence was transfected with a mixture of ten different BTV RNA transcripts using Lipofectamine 2000 reagent. At 4 h posttransfection, the medium was replaced with a 6-ml overlay consisting of minimal essential medium, 2% FBS and 1.5% (w/v) agarose. Then the plates were incubated at 35 °C in 5% CO2 for 72 h to allow plaques to appear.
Xia X., Wu W., Cui Y., Roy P, & Zhou Z.H. (2021). Bluetongue virus capsid protein VP5 perforates membranes at low endosomal pH during viral entry. Nature microbiology, 6(11), 1424-1432.
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3

BTV-10 VP6 Mutational Analysis

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BTV-10 VP6 was used for mutational analysis and reverse genetics (14 (link), 17 (link)). BTV-1 (GenBank accession numbers FJ969719 to FJ969728), the VP6 of which is fully exchangeable with that of BTV-10, was used in RCAP for viral capsids. All mutations of VP6 were generated by site-directed mutagenesis, and sequences of these mutations and primers are available upon request. Transcripts for reverse genetic analyses were prepared using the mMACHINE T7 transcription kit (Thermo); RNA transcripts for gel shifting and competition assays were prepared using T7 polymerase (Thermo), following the manufacturers’ instructions.
Sung P.Y., Vaughan R., Rahman S.K., Yi G., Kerviel A., Kao C.C, & Roy P. (2019). The Interaction of Bluetongue Virus VP6 and Genomic RNA Is Essential for Genome Packaging. Journal of Virology, 93(5), e02023-18.
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