1 of 6

Detection of Porcine (Pig) DNA in Marshmallow Products Using Real-Time PCR as the Basis for Halal Certification.

GROUP 3

Abdul Qawwiy Nasrun | Hisyam Fawwaz Giffari |

Nadira Zafira | Ratsimandresy Neçiah Fahombiazana

2 of 6

Determining the halal status of marshmallow products through pig DNA detection using Real-Time PCR techniques with the foodproof® Porcine LyoKit detection kit.

Sample Code

Product Name

Halal Label

Manufacturer

Product Form

M1

None

Yes

Brand X

Soft/Solid

M2

none

Yes

Brand X

Soft/Solid

Positive

Standard pig DNA

-

Control kit

-

Negative

Sterile water

-

Control kit

-

3 of 6

Metode

A. Ekstraksi DNA (StarPrep® Five Kit)

  • Sampel: 200 mg marshmallow dimasukkan ke tabung 2 mL
  • Tambah premix (300 µL Lysis Buffer + 500 µL Buffer M + 45 µL Proteinase K)
  • Vortex 5 detik / shaker 15 detik
  • Inkubasi 72 °C (15 menit), then inkubasi 95 °C (15 menit)
  • Sentrifugasi 13,000 × g (5 menit)
  • Supernatan digunakan (3 µL) sebagai template PCR

​

B. RT-qPCR (foodproof® Porcine LyoKit, AriaMx)

  • Pre-inkubasi: 37 °C 4 menit, then 95 °C 10 menit
  • Amplifikasi: 35 siklus (95 °C 5 s; 60 °C 60 s, data acquisition)
  • Fluorofor: FAM (DNA porcine), HEX (Internal Control)
  • Volume reaksi: 3 µL DNA + 22 µL PCR-grade water ke dalam tube kit

​

4 of 6

Hasil Data Dan Visualisasi Kurva �Amplifikasi (Simulasi)

​

​

​

​

​

​

​

  • F6 (Marshmallow 1) > Porcine No Cq; IC (HEX) 23.41 > Valid and Negatif
  • G6 (Marshmallow 2) > Porcine No Cq; IC (HEX) No Cq > Indeterminate (perlu re-uji, adanya gagal ekstraksi)

​

5 of 6

Visualisasi Kurva Amplifikasi (Simulasi)

​

​

- F6 (Marshmallow 1): kurva HEX naik melampaui threshold (IC valid), FAM datar > negatif porcine.

- G6 (Marshmallow 2): FAM dan HEX sama-sama datar > indikasi inhibisi/ekstraksi gagal (indeterminate).

6 of 6

THANK YOU