The study of transgene intergration hotspots in mammalia expression system
Mammalian expression system is very crucial for the production of proteins in numerous scientific and commercial areas as they are able to perform proper protein folding as compared to other hosts. Matrix attachment regions (MARs) are concentrated with transcription factor binding sites and have a...
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| Format: | Monograph |
| Language: | English |
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Universiti Sains Malaysia
2015
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| Online Access: | http://eprints.usm.my/59984/ http://eprints.usm.my/59984/1/PROF%20MADYA%20DR%20SHAHARUM%20SHAMSUDDIN%20-%20e.pdf |
| _version_ | 1848884319397871616 |
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| author | Shamsuddin, Shaharum |
| author_facet | Shamsuddin, Shaharum |
| author_sort | Shamsuddin, Shaharum |
| building | USM Institutional Repository |
| collection | Online Access |
| description | Mammalian expression system is very crucial for the production of proteins in numerous scientific and
commercial areas as they are able to perform proper protein folding as compared to other hosts. Matrix
attachment regions (MARs) are concentrated with transcription factor binding sites and have a strong
effect on the level of expression of transgenes, thus highly studied to increase protein production. In this
study, an expression vector carrying green fluorescent protein was transfected into mouse myeloma NSO
cell line in order to allow random integration of the vector into the host genome. Conceptually, if the
vector is integrated into a transcriptionally active site within the genome, green fluorescent protein will be
highly or constitutively expressed. Transfected cells were then subjected to sorting in order to select the
high producing cells using ClonePix FL (CPL). CPL is an automated high-throughput clone selection
instrument. DNA sequences in the genome flanking the integrated vector in the high producing cells were
subsequently retrieved using ‘genome walking’ method. Following this, the flanking regions were
sequenced and characterized using bioinformatics tools. Functional analysis was carried out by entering the
sequences into three different programs (SMARtest, MARfmder and MARscan) to determine the presence
of Matrix Attachment Regions. In addition, biochemical characterization was also done using Chromatin
Immunoprecipitation (ChIP) Assay. As a result, putative MAR elements were identified in mammalian
expression system with different lengths of MAR regions between the three algorithms. MARJnndiii proteins that were used in CMP assay were also shown to interact with the mouse |
| first_indexed | 2025-11-15T19:04:49Z |
| format | Monograph |
| id | usm-59984 |
| institution | Universiti Sains Malaysia |
| institution_category | Local University |
| language | English |
| last_indexed | 2025-11-15T19:04:49Z |
| publishDate | 2015 |
| publisher | Universiti Sains Malaysia |
| recordtype | eprints |
| repository_type | Digital Repository |
| spelling | usm-599842024-04-14T02:46:26Z http://eprints.usm.my/59984/ The study of transgene intergration hotspots in mammalia expression system Shamsuddin, Shaharum RA0421 Public health. Hygiene. Preventive Medicine RA440-440.87 Study and teaching. Research Mammalian expression system is very crucial for the production of proteins in numerous scientific and commercial areas as they are able to perform proper protein folding as compared to other hosts. Matrix attachment regions (MARs) are concentrated with transcription factor binding sites and have a strong effect on the level of expression of transgenes, thus highly studied to increase protein production. In this study, an expression vector carrying green fluorescent protein was transfected into mouse myeloma NSO cell line in order to allow random integration of the vector into the host genome. Conceptually, if the vector is integrated into a transcriptionally active site within the genome, green fluorescent protein will be highly or constitutively expressed. Transfected cells were then subjected to sorting in order to select the high producing cells using ClonePix FL (CPL). CPL is an automated high-throughput clone selection instrument. DNA sequences in the genome flanking the integrated vector in the high producing cells were subsequently retrieved using ‘genome walking’ method. Following this, the flanking regions were sequenced and characterized using bioinformatics tools. Functional analysis was carried out by entering the sequences into three different programs (SMARtest, MARfmder and MARscan) to determine the presence of Matrix Attachment Regions. In addition, biochemical characterization was also done using Chromatin Immunoprecipitation (ChIP) Assay. As a result, putative MAR elements were identified in mammalian expression system with different lengths of MAR regions between the three algorithms. MARJnndiii proteins that were used in CMP assay were also shown to interact with the mouse Universiti Sains Malaysia 2015 Monograph NonPeerReviewed application/pdf en http://eprints.usm.my/59984/1/PROF%20MADYA%20DR%20SHAHARUM%20SHAMSUDDIN%20-%20e.pdf Shamsuddin, Shaharum (2015) The study of transgene intergration hotspots in mammalia expression system. Technical Report. Universiti Sains Malaysia. (Submitted) |
| spellingShingle | RA0421 Public health. Hygiene. Preventive Medicine RA440-440.87 Study and teaching. Research Shamsuddin, Shaharum The study of transgene intergration hotspots in mammalia expression system |
| title | The study of transgene intergration hotspots
in mammalia expression system |
| title_full | The study of transgene intergration hotspots
in mammalia expression system |
| title_fullStr | The study of transgene intergration hotspots
in mammalia expression system |
| title_full_unstemmed | The study of transgene intergration hotspots
in mammalia expression system |
| title_short | The study of transgene intergration hotspots
in mammalia expression system |
| title_sort | study of transgene intergration hotspots
in mammalia expression system |
| topic | RA0421 Public health. Hygiene. Preventive Medicine RA440-440.87 Study and teaching. Research |
| url | http://eprints.usm.my/59984/ http://eprints.usm.my/59984/1/PROF%20MADYA%20DR%20SHAHARUM%20SHAMSUDDIN%20-%20e.pdf |