Microscopic detection of chloroplast transgenic plastids using fluorescent probe
Background: Fluorescent marker genes have modernized many areas of molecular biology, specifically in plant biotechnology and genetic engineering studies. The use of fluorescent proteins permits the scientists to purify the desired clones visually in transformation work. Green fluorescent protein (g...
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| Format: | Article |
| Language: | English |
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The Running Line
2018-05-01
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| Series: | Advancements in Life Sciences |
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| Online Access: | http://www.als-journal.com/536-18/ |
| _version_ | 1827853852538306560 |
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| author | Shahid Nazir Muhammad Sarwar Khan |
| author_facet | Shahid Nazir Muhammad Sarwar Khan |
| author_sort | Shahid Nazir |
| collection | DOAJ |
| description | Background: Fluorescent marker genes have modernized many areas of molecular biology, specifically in plant biotechnology and genetic engineering studies. The use of fluorescent proteins permits the scientists to purify the desired clones visually in transformation work. Green fluorescent protein (gfp) derived from Aequorea victoria has been the most common and favorite fluorescent marker which is being widely used as a visual selection marker gene. It can be easily visualized under UV light without the involvement of any substrate and is non-destructive as well.
Method: A species-specific chloroplast transformation vector was constructed with gfp as a fluorescent marker gene. The recombinant vector was biolistically integrated in tobacco plastome and transgenic cells were initially screened on spectinomycin containing regeneration medium.
Results: The successful plastome integration was verified by using cellular DNA from drug resistant clones in PCR and southern blotting. The expression of gfp in transplastomic clones was microscopically investigated using simple florescent as well as confocal laser scanning microscopes.
Conclusion: Regeneration of transgenic plants was significantly helped by visual identification of fluorescent at different stages of development, also enabling to identify the homozygous and heterozygous tissues. No toxic effect of the gfp was observed and lack of toxicity as maintained by normal phenotypic performance of plants. |
| first_indexed | 2024-03-12T11:11:49Z |
| format | Article |
| id | doaj.art-9e4ad7deed0e46c0b339c7ec365cc570 |
| institution | Directory Open Access Journal |
| issn | 2310-5380 2310-5380 |
| language | English |
| last_indexed | 2024-03-12T11:11:49Z |
| publishDate | 2018-05-01 |
| publisher | The Running Line |
| record_format | Article |
| series | Advancements in Life Sciences |
| spelling | doaj.art-9e4ad7deed0e46c0b339c7ec365cc5702023-09-02T02:44:14ZengThe Running LineAdvancements in Life Sciences2310-53802310-53802018-05-0153121129Microscopic detection of chloroplast transgenic plastids using fluorescent probeShahid Nazir0 Muhammad Sarwar Khan1Agricultural Biotechnology Research Institute, Faisalabad , PakistanCenter of Agricultural Biochemistry & Biotechnology (CABB), University of Agriculture, Faisalabad, PakistanBackground: Fluorescent marker genes have modernized many areas of molecular biology, specifically in plant biotechnology and genetic engineering studies. The use of fluorescent proteins permits the scientists to purify the desired clones visually in transformation work. Green fluorescent protein (gfp) derived from Aequorea victoria has been the most common and favorite fluorescent marker which is being widely used as a visual selection marker gene. It can be easily visualized under UV light without the involvement of any substrate and is non-destructive as well. Method: A species-specific chloroplast transformation vector was constructed with gfp as a fluorescent marker gene. The recombinant vector was biolistically integrated in tobacco plastome and transgenic cells were initially screened on spectinomycin containing regeneration medium. Results: The successful plastome integration was verified by using cellular DNA from drug resistant clones in PCR and southern blotting. The expression of gfp in transplastomic clones was microscopically investigated using simple florescent as well as confocal laser scanning microscopes. Conclusion: Regeneration of transgenic plants was significantly helped by visual identification of fluorescent at different stages of development, also enabling to identify the homozygous and heterozygous tissues. No toxic effect of the gfp was observed and lack of toxicity as maintained by normal phenotypic performance of plants.http://www.als-journal.com/536-18/Fluorescent markerMicroscopyChloroplast transformationTobacco |
| spellingShingle | Shahid Nazir Muhammad Sarwar Khan Microscopic detection of chloroplast transgenic plastids using fluorescent probe Advancements in Life Sciences Fluorescent marker Microscopy Chloroplast transformation Tobacco |
| title | Microscopic detection of chloroplast transgenic plastids using fluorescent probe |
| title_full | Microscopic detection of chloroplast transgenic plastids using fluorescent probe |
| title_fullStr | Microscopic detection of chloroplast transgenic plastids using fluorescent probe |
| title_full_unstemmed | Microscopic detection of chloroplast transgenic plastids using fluorescent probe |
| title_short | Microscopic detection of chloroplast transgenic plastids using fluorescent probe |
| title_sort | microscopic detection of chloroplast transgenic plastids using fluorescent probe |
| topic | Fluorescent marker Microscopy Chloroplast transformation Tobacco |
| url | http://www.als-journal.com/536-18/ |
| work_keys_str_mv | AT shahidnazir microscopicdetectionofchloroplasttransgenicplastidsusingfluorescentprobe AT muhammadsarwarkhan microscopicdetectionofchloroplasttransgenicplastidsusingfluorescentprobe |