GEL ELECTROPHORESIS

Visualisasi hasil PCR

Horizontal Electrophoresis:
- Agarose gel
-TBE or TAE buffer
- Ethidium bromide or SYBR Green staining
- Loading dye

Vertical Electrophoresis:
- Agarose gel
-TBE buffer
- Ethidium bromide or SYBR Green staining
- Loading dye

Agarose gel concentration

100 – 700 bp ………..2 % (ultra or high grade agarose >> more than 2%)
500 – 1500 bp………1.5 %
1000 – 5000 bp………1 % or 0.7 %
 4

6
3
2

1
5
 Question
1. Why the El-fo running from the negative
catode
2. EtBr (0,5µL/L) how many EtBr in 100 mL TEA
1x?
Resolution of DNA in Agarose
Agarose Effective separation range
Consentration (%) (Kbp Linear)
2.0 0.1-3
1.5 0.2-4
1.2 0.4-6
1.0 0.5-7
0.8 0.7-10
0.6 1-20
0.3 5-60
 Elektroforesis ; Pembuatan Agar (Agarose)

Menimbang agarose mendidihkan

dan mengukur buffer Mempersiapkan tray

Dituang ke tray
setelah ditambah
ethidium bromida

Siap digunakan
Elektroforesis Horizontal

Kira-kira 5 uL ditambah 2
uL loading dye
dimasukkan pada tiap
sumur gel

Gel dimasukkan ke chamber

Tambahkan larutan buffer

Elektroforesis sedang berjalan Diberi aliran listrik

Vertical Electrophoresis
Note: DNA is negatively charged
Visualisasi

Gel diletakkan diatas UV box UV dinyalakan

Diatur fokus dan dilakukan

pemotretan
- UV box
- UV – Polaroid film
- GelDoc
Allele Specific PCR (AS-PCR)

Alel 1 TTGCTGGGAGCTGTTCTACTGCTATCAGCTCTGCCCGGTCATGACCAGGAAA
Alel 2 TTGCTGGGAGCTGTTCTACTGCTATTAGCTCTGCCCGGTCATGACCAGGAAA
Primer 1 TGTTCTACTGCTATC TGGTCCTTT
Primer 2 TGTTCTACTGCTATT

M 1 2 1 2 1 2

S1 S2 S3
Alel1 Alel1/2 Alel2