000			03690nas|a2200433 i 4500
005			20260817172938.0
008			020717s2002 th u m rtt 00| a1eng d
035			_a.b11861393
099		9	_aAIT Thesis no. BP-02-01
100	1		_aDar, Shabir Ahmad
245	1	0	_aCloning of the chitin deacetylase gene from the fungus Mucor rouxii into the yeast Pichia pastoris
260			_aBangkok : _bAsian Institute of Technology, _c2002
300			_a61, [12] leaves
490	1		_aThesis ; _vno. BP-02-01
502			_aThesis (M.Sc.) - Asian Institute of Technology
500			_aA thesis submitted in partial fulfillment of the requirement for the degree of Master of Science, School of Environment, Resources and Development
502			_aThesis (M. Sc.) - Asian Institute of Technology, 2002
520			_aChitin, which occurs in the exoskeleton of arthropods and in the cell wall of fungi, is an abundant renewable natural resource. Chitosan, the deacetylated form of chitin has a broad variety of industrial and biomedical applications. It is traditionally produced by a harsh thermochemical procedure that is environmentally unsafe and not easily controlled. The use of chitin deacetylases offers the possibility of an alternative procedure to the chemical process for the deacetylation of chitin. Molecular cloning of CDA gene into a suitable host is therefore explored to achieve efficient production of the enzyme. This work aimed at the cloning of CDA gene from Mucor rouxii and the constructing of CDA-overexpressing strains of Pichia pastoris. Chitin deacetylase (CDA) gene after amplification from Mucor rouxii ATCC 24905 was cloned into two of the E.coli-Pichia pastoris shuttle vectors, pPIC3.5K and pPIC9K. In case of pPIC9K the gene was cloned in phase with the a-secretion signal factor present in the parent vector for the secreted expression of CDA. The gene was put in both vectors under the transcriptional control of Pichia pastoris alcohol oxidase gene promoter. The gene was transformed into Pichia pastoris GS115 (methanol utilization fast, Mut) to obtain overexpressing strains. Pichia transformants were selected on histidine deficient medium and screened for their ability to use methanol as a sole source of carbon. The selected clones were screened for transformants resistant to G418 (Geneticin). Integration of the gene in Pichia genome was confirmed through PCR analysis of the transformants. The intracellular and secreted expression of the cloned CDA gene in Pichia transformants was analyzed through SDSp AGE analysis.
650	1	0	_aMolecular cloning
650	1	0	_aChitin
700	1		_aStevens, Willem Frans, _eChairperson
700	1		_aHegarat, Francoise Le, _eExamination Committee
700	0		_aSuwalee Chandrkrachang, _eExamination Committee
700	0		_aSunee Nitisinprasert, _eExamination Committee
700	1		_aMacaranas, Julie, _eExamination Committee
710	2		_aThe Royal Netherlands Government, _eScholarship Donor
810	2		_aAsian Institute of Technology. _tThesis ; _vno. BP-02-01
856			_3Full-Text _uhttp://203.159.5.9/ait-thesis/detail.php?q=B08077
907			_a.b11861393 _bmnait _cu
902			_a240402
998			_b0 _c020717 _dm _ea _fu _g0
945			_lmnait
945			_lmnarc
942			_c22
942			_c40
909			_aBarcode : 30050120682959 _bCREATED : 2013-06-09 _cRECORD # : i12739972 _dLPATRON : 0 _eLCHKIN : - _f# RENEWALS : 0 _g# OVERDUE : 0 _hIUSE3 : 0 _iTOT CHKOUT : 0 _jTOT RENEW : 0
909			_aBarcode : 30050120350946 _bCREATED : 2016-10-06 _cRECORD # : i13029502 _dLPATRON : 0 _eLCHKIN : - _f# RENEWALS : 0 _g# OVERDUE : 0 _hIUSE3 : 0 _iTOT CHKOUT : 0 _jTOT RENEW : 0
999			_c18116 _d18116