Yıl 2018, Cilt 22, Sayı 6, Sayfalar 1508 - 1517 2018-12-01

Cloning, Expression and Characterization of Xylanase (xyn-akky1) from Bacillus subtilis in Escherichia coli
Bacillus subtilis’e Ait Ksilanaz (xyn-akky1)’in Escherichia coli’de Klonlanması, Ekspresyonu ve Karakterizasyonu

sema bilgin [1] , Yakup ULUSU [2] , Hülya KUDUĞ [3] , İsa GÖKÇE [4]

218 503

In this study Bacillus subtilis akky1 strain was isolated from the soil of beech forest in Akkuş City, Ordu Province, Turkey. The identification of the strain akky1 done by PCR amplification 16S rRNA. The full-length 16S rRNA sequence showed the highest nucleotide similarity (100%) with Bacillus subtilis strain B7 (KC310823.1). Spesific oligonucleotides targeting the sequence of Bacillus subtilis xylanase gene given in GenBank were used to amplify a 642-bp fragment from genomic DNA. The gene encoding xylanase was cloned into pET28b (+) plasmid vector, sequenced and expressed in Escherichia coli BL21 (DE3). The hexahistidine (6xHis) tagged fusion protein was purified using nickel affinity chromatography and the xylanase activity was measured. The molecular mass of the purified xylanase was approximately 26 kDa as estimated by SDS-PAGE. The xylanase had optimal activity at pH 6.0 and 60°C. The Km values of the recombinant enzyme towards beechwood was 3,33 mg/ml.

 Bu çalışmada, Bacillus subtilis akky1 straini Türkiye’nin Ordu İli’nin Akkuş ilçesindeki kayın ormanından alınan topraktan izole edilmiştir. akky1 straini 16S rRNA analizi ile tanımlanmıştır. 16S rRNA dizisi Bacillus subtilis strain B7 (KC310823.1) ile %100’lük bir benzerlik göstermiştir. GenBank’da verilen Bacillus subtilis ksilanazına ait gen dizisi esas alınarak tasarlanan primerler kullanılarak genomic DNA’dan 642 bp’lik bir DNA fragmenti elde edilmiştir. Ksilanazı kodlayan gen, pET28b (+) ekspresyon vektörüne klonlanmış ve Escherichia coli BL21 (DE3)’de ekpresse edilmiştir. Rekombinant olarak üretilen protein nikel afinite kromatografisi ile saflaştırılmış ve ksilanaz aktivitesi belirlenmiştir. Saflaştırılan ksilanazın moleküler ağırlığı SDS-PAGE ile 26.0 kDa olarak belirlenmiştir. Rekombinant enzimin optimum pH’sı 6.0 ve sıcaklığı 60°C, Km değeri ise 3.33 mg/ml olarak tespit edilmiştir.  

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Birincil Dil en
Konular Mühendislik, Kimya, Biyoloji, Temel Bilimler
Yayımlanma Tarihi December 2018
Dergi Bölümü Araştırma Makalesi
Yazarlar

Yazar: sema bilgin
Kurum: GAZİOSMANPAŞA ÜNİVERSİTESİ, FEN-EDEBİYAT FAKÜLTESİ, KİMYA BÖLÜMÜ
Ülke: Turkey


Yazar: Yakup ULUSU
Kurum: KARAMANOĞLU MEHMETBEY ÜNİVERSİTESİ, MÜHENDİSLİK FAKÜLTESİ, BİYOMÜHENDİSLİK BÖLÜMÜ
Ülke: Turkey


Yazar: Hülya KUDUĞ
Kurum: GAZİOSMANPAŞA ÜNİVERSİTESİ, MÜHENDİSLİK VE DOĞA BİLİMLERİ FAKÜLTESİ
Ülke: Turkey


Yazar: İsa GÖKÇE
Kurum: GAZİOSMANPAŞA ÜNİVERSİTESİ, MÜHENDİSLİK VE DOĞA BİLİMLERİ FAKÜLTESİ
Ülke: Turkey


Bibtex @araştırma makalesi { saufenbilder327153, journal = {Sakarya University Journal of Science}, issn = {1301-4048}, eissn = {2147-835X}, address = {Sakarya Üniversitesi}, year = {2018}, volume = {22}, pages = {1508 - 1517}, doi = {10.16984/saufenbilder.327153}, title = {Cloning, Expression and Characterization of Xylanase (xyn-akky1) from Bacillus subtilis in Escherichia coli}, key = {cite}, author = {GÖKÇE, İsa and bilgin, sema and ULUSU, Yakup and KUDUĞ, Hülya} }
APA bilgin, s , ULUSU, Y , KUDUĞ, H , GÖKÇE, İ . (2018). Cloning, Expression and Characterization of Xylanase (xyn-akky1) from Bacillus subtilis in Escherichia coli. Sakarya University Journal of Science, 22 (6), 1508-1517. DOI: 10.16984/saufenbilder.327153
MLA bilgin, s , ULUSU, Y , KUDUĞ, H , GÖKÇE, İ . "Cloning, Expression and Characterization of Xylanase (xyn-akky1) from Bacillus subtilis in Escherichia coli". Sakarya University Journal of Science 22 (2018): 1508-1517 <http://www.saujs.sakarya.edu.tr/issue/31266/327153>
Chicago bilgin, s , ULUSU, Y , KUDUĞ, H , GÖKÇE, İ . "Cloning, Expression and Characterization of Xylanase (xyn-akky1) from Bacillus subtilis in Escherichia coli". Sakarya University Journal of Science 22 (2018): 1508-1517
RIS TY - JOUR T1 - Cloning, Expression and Characterization of Xylanase (xyn-akky1) from Bacillus subtilis in Escherichia coli AU - sema bilgin , Yakup ULUSU , Hülya KUDUĞ , İsa GÖKÇE Y1 - 2018 PY - 2018 N1 - doi: 10.16984/saufenbilder.327153 DO - 10.16984/saufenbilder.327153 T2 - Sakarya University Journal of Science JF - Journal JO - JOR SP - 1508 EP - 1517 VL - 22 IS - 6 SN - 1301-4048-2147-835X M3 - doi: 10.16984/saufenbilder.327153 UR - http://dx.doi.org/10.16984/saufenbilder.327153 Y2 - 2017 ER -
EndNote %0 Sakarya University Journal of Science Cloning, Expression and Characterization of Xylanase (xyn-akky1) from Bacillus subtilis in Escherichia coli %A sema bilgin , Yakup ULUSU , Hülya KUDUĞ , İsa GÖKÇE %T Cloning, Expression and Characterization of Xylanase (xyn-akky1) from Bacillus subtilis in Escherichia coli %D 2018 %J Sakarya University Journal of Science %P 1301-4048-2147-835X %V 22 %N 6 %R doi: 10.16984/saufenbilder.327153 %U 10.16984/saufenbilder.327153
ISNAD bilgin, sema , ULUSU, Yakup , KUDUĞ, Hülya , GÖKÇE, İsa . "Bacillus subtilis’e Ait Ksilanaz (xyn-akky1)’in Escherichia coli’de Klonlanması, Ekspresyonu ve Karakterizasyonu". Sakarya University Journal of Science 22 / 6 (Aralık 2018): 1508-1517. http://dx.doi.org/10.16984/saufenbilder.327153