Professional Documents
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Anonim.2006.Fagosistosis.http://pkukmweb.ukm.my/~danial/Fagositosis.ht
ml. Diakses 7 November 2008.
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Williams, H., Shaw, M.A. 1995. Genomic organization and
Sequence of the Human NRAMP Gene: Identification and Mapping
of a promoter Region Polymorphism. Molec. Med. 1: 194-205.
Bellamy, R., Ruwende, C., Corrah, T., McAdam, K. P. W. J., Whittle, H.C.,
Hill, A.V.S. 1998. Variations in the NRAMP1 gene and susceptibility
to tuberculosis in West Africans. New Eng. J. Med. 338: 640-644.
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resistance-associated macrophage prote in: cDNA cloning,
chromosomal mapping, genomic organization, and tissue-specific
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Darm owandowo W. Demam Tifoid. Dalam : Soedarmo SS, Garna H,
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of Severe Typhoid Fever. Trends in Microbiology. Vol.9 No.7.
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Sampel darah
Medium Enrichment
(3 ml darah + 9 ml Ox Bile Broth)
Medium selektif
(1 ml dari medium pengaya + 15 ml SSA)
RFLP-PCR
Pengamatan
intensitas warna
Elektroforesis gel
Pengamatan
Analisis Data
Lampiran 3 . Skema Kerja Ekstraksi DNA
Endapan + 1 ml Aseton
(dihomogenkan, disentrifugasi 12.000 rpm, 30 detik)
Endapan + Buffer TE 90 µL
(divortex hingga homogen per sampel)
Diambil 5 µl + serum
penderita, dihomogenkan
PCR
PCR mix :
2,5 µl 10X buffer PCR + 0,5 µl 10X Ex Taq Polimerase + 2 µl
MgCl2+ 1 µl dNTPs + 1 µl Primer For + 1 µl Primer Rev + 2,5
µl Sampel ekstraksi DNA + 13 µl H2O
Amplifikasi 35 siklus
- Denaturasi 94 oC 45 detik
- Annealing 57 oC 45 detik
o
- Elongasi 72 C 45 detik
Produk PCR
PCR-RFLP
(campuran 4 µl DNA hasil PCR + 16 µl campuran enzim restriksi terdiri
dari : 0,5 enzim restriksi Ava II dan 0,5 µl NEB buffer serta 7 µl
aquades. Campuran diiinkubasi pada suhu 37 oC, 1x24 jam diruangan)
Pengamatan pada UV
Transilluminator
Lampiran 6. Sequence Nukleotida Gen NRAMP-1, Posisi Primer dan
Letak Pemotongan oleh Enzim Restriksi
NO UMUR SEX LAMA DEMAM NO.DARAH HASIL KULTUR HASIL DIPSTIK SUMBER
NO UMUR SEX LAMA DEMAM NO.DARAH HASIL KULTUR HASIL DIPSTIK SUMBER