You are here

Home » Content

HBeAg gen bölgesinin Saccharomyces cerevisiae’da ekspresyonu

Expression of HBeAg gene region in Saccharomyces cerevisiae

Journal Name:

  • Gülhane Tıp Dergisi

Publication Year:

  • 2009

Key Words:

Keywords (Original Language):

Author Name
Abstract (2. Language): 
HBeAg/anti-HBe seroconversion is one of the markers frequently used in the diagnosis of hepatitis B infection. In this study, the “e” antigen gene region of the hepatitis B virus (HBV) was transformed to Saccharomyces cerevisiae and expressed by the yeast in order to obtain HBeAg protein used in serologic tests. Firstly, HBeAg gene region was amplified by PCR method. Later, the amplicons were cloned into the plasmid pYES2.1 via the TOPO® TA expression kit and transformed into the competent bacteria (TOPO10F’ Escherichia coli). The plasmid pYES2.1+HBeAg was isolated from the competent bacteria, which was cultivated on LB media supplemented with ampicillin, and transformed to Saccharomyces cerevisiae via the S.c. EasyComp Transformation Kit. Finally, whether HBeAg was expressed by the yeast was checked through two different automatized systems. HbeAg producing yeast cells obtained may render it possible to purely isolate proteins in future studies.
Bookmark/Search this post with
Abstract (Original Language): 
HBeAg/anti-HBe serokonversiyonu hepatit B enfeksiyonlarının tanısında sık kullanılan göstergelerden birisidir. Bu çalışmada serolojik testlerde kullanılan HBeAg proteinini elde etmek amacıyla hepatit B virüsü (HBV) “e” antijen gen bölgesi Saccharomyces cerevisiae’ya aktarıldı ve eksprese ettirildi. Bunun için öncelikle HBeAg gen bölgesi PCR yöntemiyle çoğaltıldı. Daha sonra amplikonlar TOPO® TA ekspresyon kiti ile pYES2.1 plazmidine klonlandı ve kompetan bakteriye (TOPO 10F’ Escherichia coli) transforme edildi. Ampisilinli LB agarda çoğaltılan kompetan bakteriden pYES2.1+HBeAg plazmidi izole edildi ve S.c. EasyComp transformasyon kitiyle Saccharomyces cerevisiae’ya transforme edildi. Son olarak, HBeAg’nin maya tarafından eksprese edilip edilmediği iki farklı otomatize sistem aracılığıyla kontrol edildi. Elde edilen HBeAg üreten maya hücreleri sonraki çalışmalarda üretilen proteinlerin pür olarak elde edilmesine imkan sağlayabilir.
FULL TEXT (PDF): 
PDF icon arastirmax-hbeag-gen-bolgesinin-saccharomyces-cerevisiaeda-ekspresyonu.pdf
  • 2
91-93
  • Turkish

REFERENCES

References: 

Kaynaklar
1. Badur S. Hepatit A, B ve D virusları. In: Ustaçelebi Ş,
Abacıoğlu H, Badur S (eds). Moleküler, Klinik ve Tanısal
Viroloji. Ankara: Güneş Kitabevi, 2004: 175-202.
2. Harrison TJ, Dusheiko GM, Zuckerman AJ. Hepatitis
viruses. In: Zuckerman AJ, Banatvala JE, Schoub BD,
Griffiths PD, Mortimer P (eds). Principles and Practice of
Clinical Virology. 6th ed. London: John Wiley & Sons
Ltd, 2009: 273-319.
3. Seeger C, Zoulim F, Mason WS. Hepadnaviruses. In: Knipe
DM, Howley PM (eds). Fields Virology. 5th ed. New York:
Lippincott Williams & Wilkins, 2007: 1253-1304.
4. Yapar M, Aydogan H, Pahsa A, et al. Rapid and quantitative
detection of Crimean-Congo Hemorragic Fever Virus by
one step real-time reverse transcriptase PCR. Jpn J Infect
Dis 2005; 58: 358-362.
5. Yapar M, Guney C, Basustaoglu AC, Kubar A. Expression
of hepatitis B “e” antigen gene in Escherichia coli.
Mikrobiyol Bul 2001; 35: 273-278.
6. h t t p : / / s t r u c t u r e . b i o c h e m . q u e e n s u . c a / p r o t o c o l s /
compcells.htm (Son erişim tarihi: 17.03.2008)
7. pYES2.1 TOPO® TA Expression Kit. Five-minute cloning
of Taq polymerase-amplified PCR products for regulated
expression in Saccharomyces cerevisiae. Catalog no.
K4150-01. Invitrogen. User manual.
8. Guthrie CC, Fink GR. Guide to yeast genetics and
molecular biology. In: Abelson JN, Simon MI (eds).
Methods in Enzymology. Vol. 194. San Diego: Academic
Press, 1991.
9. Saracli MA, Sener K, Gonlum A, Yildiran ST, Wickes
BL. Genotyping of clinical Rhodotorula mucilaginosa
isolates by pulsed field gel electrophoresis. Mycoses
2003; 46: 487-491.
10. Kniskern PJ, Hagopian A, Montogomery DL, et al.
Unusually high level expression of foreign gene (hepatitis
B virus core antigen in Saccharomyces cerevisiae). Gene
1986; 46: 135-141.
11. Kubar A, Yapar M, Ozyurt M, Haznedaroglu T, Gun
H. Cloning of hepatitis B virus surface gene region to
Escherichia coli. Flora 1988; 3: 183-186.
12. Yapar M, Guney C, Saracli MA, Kilic A, Kubar A.
Expression of HBsAg gene in insect (Trichiplusia ni) cell
culture. Flora 2001; 6: 108-113.

Thank you for copying data from http://www.arastirmax.com