rt-pcr mediated cloning of human growth hormone gene and i ts expression in e. coli

نویسندگان

m azarnoosh from the department of biotechnology, pasteur institute of iran, tehran, islamic republic of iran.

s zeinali

sm tabatabaei

a ziaee

چکیده

human growth hormone (hgh) genomic sequence containing 5 exons and 4 introns was cloned in pcdna-3 and the constructed plasmid was subsequently used for transfection ofnlli-3t3 cell line using lipofection technique. expression of hgh in stably transfected cells was assayed using elisa. total rna was extracted from transfected cells and hgh cdna was amplified by rt-pcr using specific primers. the product thus obtained was cloned in pgem-t vector and the presence of the growth hormone coding region was verified by restriction enzyme analysis and southern blotting. the hgh cdna fragment was cloned in pqe-30 and the expression of hgh gene in e. coli was assayed using elisa and immunoblotting. in this experiment 20.9 µg/ml purified rhgh was obtained.

برای دانلود باید عضویت طلایی داشته باشید

برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید

اگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید

منابع مشابه

RT-PCR MEDIATED CLONING OF HUMAN GROWTH HORMONE GENE AND I TS EXPRESSION IN E. coli

Human growth hormone (hGH) genomic sequence containing 5 exons and 4 introns was cloned in pcDNA-3 and the constructed plasmid was subsequently used for transfection ofNlli-3T3 cell line using lipofection technique. Expression of hGH in stably transfected cells was assayed using ELISA. Total RNA was extracted from transfected cells and hGH cDNA was amplified by RT-PCR using specific primers...

متن کامل

Cloning and Expression of Bst DNA Polymerase I Gene in E. coli BL21‎

خلاصه DNA پلیمرازها علاوه بر کاربردشان در همسانه سازی و تصحیح ، در انواع تکنیک های ملکولی مانند تکثیر DNA ، جهش های نقطه ای ، توالی یابی DNA ، انواع مختلف PCR ، LAMP و ...اهمیت دارند. پس از کشف PCR تلاش هایی مبنی بر تشخیص و جداسازی آنزیم های مقاوم به دماهای بالا که توانایی تکثیر موثر DNA در دماهای بالا انجام گرفت. در این پژوهش ، سویه Geobacillus stearothermophilus strain 10 به منظور...

متن کامل

CLONING AND EXPRESSION OF A HUMAN INTERFERON a2 GENE IN E. COLI

The plasmid pALCA1SIFN containing cDNA that encodes the human interferon a-2b was obtained from the ATCC(no. 531667). In this system the expression of the gene is under the control of an alcA promoter. alcA p is a specific promoter for expression of different genes in Aspergillusfilamentous. In this plasmid the coding region of IFN?-2b is preceded by the coding region of a synthetic signal ...

متن کامل

cloning and expression of human vasohibin1 gene in e. coli

background: angiogenesis is an important process in various physiologic and pathologic states. the most significant stimulator of angiogenesis is vascular endothelial growth factor (vegf). in contrast, vasohibin1 acts as an angiogenesis inhibitor which specifically inhibits new vessels formation. the aim of the present study was cloning and expression of vasohibin1 gene in e. coli as well as pu...

متن کامل

PCR-mediated Expression of the Human GM-CSF Gene in Escherichia coli

Four exons of the human genomic GM-CSF gene were assembled together using gene splicing by overlap extension (SOE) method. The resulting nucleotide sequence was cloned in the pET23a(+) ex‌pression vector under the control of strong bacteriophage T7 transcription and translation signals. The construct obtained was Transferred into the E. coli strain, BL21(DE3) pLysS and IPTG was used for inducti...

متن کامل

منابع من

با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید


عنوان ژورنال:
medical journal of islamic republic of iran

جلد ۱۲، شماره ۳، صفحات ۲۸۵-۲۹۰

کلمات کلیدی

میزبانی شده توسط پلتفرم ابری doprax.com

copyright © 2015-2023