Difference between revisions of "Part:BBa K805012"
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We construct the gene on the carrier displaying on the surface of Pichia pastoris and express it it in Pichia pastoris with electroporation. Then we detect the expression on the surface of Pichia pastoris by observing with fluorescent microscope and by flow cytometry analysis. | We construct the gene on the carrier displaying on the surface of Pichia pastoris and express it it in Pichia pastoris with electroporation. Then we detect the expression on the surface of Pichia pastoris by observing with fluorescent microscope and by flow cytometry analysis. | ||
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[[Image:Luorescent_microscope_6.jpg]] | [[Image:Luorescent_microscope_6.jpg]] | ||
| + | We fused FLAG-tag with N-terminal of Ruxyn gene on Ruxyn-pPIC9k vector. Label cells with Rabbit Anti Flag-Tag Polyclonal antibody and Goat Anti Rabbit RPE Polyclonal antibody. Result of flow cytometry analysis is as follows: fluorescence intensity x-mean value of Pichia pastoris with pPIC9k empty vector is 1.14, with Ruxyn-pPIC9k is 8.56. | ||
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| + | [[Image:Hust_part5.jpg]] | ||
| + | [[Image:Hust_part6.jpg]] | ||
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| + | Left: pPIC9k negative control (x-mean: 1.14/13.4); Right: Ruxyn (x-mean: 8.56/19.9) | ||
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Latest revision as of 04:47, 27 September 2012
Ruxyn1 Glycosyl hydrolase (GH) family 43 b-Dxylosidase/a-L-arabinofuranosidase Ruxyn can hydrolyze p-nitrophenyl-b-D-xylopyranoside (pNPX), p-nitrophenyla-L-arabinofuranoside (pNPA), and xylo-oligosaccharide substrates. Ruxyn shows high synergism with endoxylanase,greatly elevating the reducing sugars released from brichwood xylans, and converted most intermediate xylo-oligosaccharide hydrolysate into xylose.
We construct the gene on the carrier displaying on the surface of Pichia pastoris and express it it in Pichia pastoris with electroporation. Then we detect the expression on the surface of Pichia pastoris by observing with fluorescent microscope and by flow cytometry analysis.
We fused FLAG-tag with N-terminal of Ruxyn gene on Ruxyn-pPIC9k vector. Label cells with Rabbit Anti Flag-Tag Polyclonal antibody and Goat Anti Rabbit RPE Polyclonal antibody. Result of flow cytometry analysis is as follows: fluorescence intensity x-mean value of Pichia pastoris with pPIC9k empty vector is 1.14, with Ruxyn-pPIC9k is 8.56.
Left: pPIC9k negative control (x-mean: 1.14/13.4); Right: Ruxyn (x-mean: 8.56/19.9)
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 396
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 373
- 1000COMPATIBLE WITH RFC[1000]