UNIVERSITAS AIRLANGGA



Detail Article

Folia Medica Indonesiana

ISSN 0303-7932

Vol. 49 / No. 2 / Published : 2013-04

Order : 7, and page :101 - 108

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Original Article :

Direct cloning of melanoma antigen 1 (mage-1) and the e2 gene of hog cholera virus (hcv) from the blunt end pcr product using pet 101/d-topo

Author :

  1. Gondo Mastutik*1
  2. Rosmelati Situmeang*2
  1. Dosen Fakultas Kedokteran
  2. Technical Services Unit Area, Veterinary Public Health Laboratory, East Kalimantan

Abstract :

The objective of this study was to clone the full length of coding region of Mage-1 and E2 gene of Hog Cholera Virus (HCV) usingthe plasmid pET101/D-TOPO to get the corresponding recombinant plasmid. Samples were obtained from the liver tissue withhepatocellular carcinoma and a pig spleen infected by HCV from Sukoharjo Middle Java. cDNA were amplified by PCR usingGMTOPOF-GMTOPOR for Mage-1 and RS-1-RS-2 for E2 HCV.The purified PCR products were cloned into pET101/D-TOPO asdirect cloning expression vector. The gene target was transformed into E. coli Top10. Analysis of the recombinant plasmid wasundertaken by sequencing and restriction test. The PCR of Mage-1 gene resulted fragments at + 1105bp for first round PCR and at+ 931bp for the second round. The PCR of E2 gene HCV produced fragment at + 1200 bp. The DNA targets were cloned intopET101/D-TOPO from the blunt end PCR product directly. The sequence of full length of coding region of Mage-1 contained 927nucleotides that encoded 309 amino acids residues. The sequence of the full length coding region of E2 gene HCV was encoded by1218 nucleotide. EcoRV enzyme cuts vector pET101/D-TOPO at nucleotide positions at 545 and 4775bp. The result of EcoRVrestriction had band at 4230 and 2450 for pETGM/MAGE1-HCC and 4200 and 2700bp for pETRS/E2-SH. The recombinant plasmidpETGM/MAGE1-HCC and pETRS/E2-SH were obtained from the blunt end PCR product for direct cloning using pET101/D-TOPO.(FMI 2013;49:101-108)

Keyword :

direct cloning, Mage-1, E2 Hog Cholera Virus, pET101/D-TOPO,


References :

Casali N and Preston A,(2003) E. coli Plasmid Vectors: Methods and Applications Totowa : Humana Press

Mastutik G, Hardjowijoto S, Lunardi JH, Puspaningsih NNT, Soetjipto, Kusumobroto HO, Putra ST,(2008) Molecular analysis of the coding sequence of mage-1 gene fromhepatocelular carcinoma patient and testis - : Folia Medica Indonesiana





Archive Article

Cover Media Content

Volume : 49 / No. : 2 / Pub. : 2013-04
  1. The Effectiveness Of Topical Garlic Extract For Methicillin Resistant Staphylococcus Aureus Treatment In Burns
  2. Mycobacterium Tuberculosis Immunoglobulin M (igm) And Immunoglobulin G (igg) Survey In Hiv-infected Donors
  3. Comparing The Effect Of Red Yeast Rice, Date Palm, And Guava Leaf Extract On Thrombocyte And Megakaryocyte Count In Thrombocytopenic White Rats
  4. Maternally Inherited Nucleotide Variation Of Mitochondrial Dna (mtdna) D-loop Region 126 Pb (nt: 34-159 Hvs Ii) In Madurese Individuals
  5. Rapid Culture Method Using Biphasic Media On Bacteriologic Diagnosis To Detect Mycobacterium Tuberculosis For Determining Pulmonary Tuberculosis
  6. Effect Of Carbide And Cacl2 Concentration On Vitamin C Contents Of Banana Kepok
  7. Direct Cloning Of Melanoma Antigen 1 (mage-1) And The E2 Gene Of Hog Cholera Virus (hcv) From The Blunt End Pcr Product Using Pet 101/d-topo
  8. Quantitation Of Microorganisms In Lower Respiratory Tract Secretions
  9. Bmd-qus Assessment Result Appropriates To Diagnose Osteoporosis
  10. Acid-fast Bacteria M. Leprae Examination In Leprosy Patients Before And After Treatment
  11. Diagnostic Role Of Intra Operative Ovarian Cyst Fluid Cytology In Determining Malignancy
  12. Correlation Between Adolescents' Knowledge On Drugs Hazard And Their Behavior In Drug Abuse In Tulungagung
  13. Basal Insulin As Add-on Therapy To Oral Glucose-lowering Drugs In Insulin Naive Patients With Type 2 Diabetes Mellitus In Primary Public Health Center

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