ANALISIS IN SILICO PRIMER MULTIPLEX GEN HOUSEKEEPING PADA IKAN SIDAT (Anguilla bicolor)

Yusi Yustami, - (2021) ANALISIS IN SILICO PRIMER MULTIPLEX GEN HOUSEKEEPING PADA IKAN SIDAT (Anguilla bicolor). S1 thesis, Universitas Pendidikan Indonesia.

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Abstract

Indonesia merupakan salah satu negara sebagai pengekspor ikan sidat (Anguilla bicolor), produk dihasilkan melalui budidaya benih (glass eel) yang ditangkap di muara sungai. Budidaya pemijahan ikan sidat sulit dilakukan, dapat disebabkan kondisi lingkungan yang mengganggu sistem metabolisme tubuh ikan. Berdasarkan hal tersebut tujuan penelitian ini adalah memonitoring kelulushidupan ikan sidat dengan cara memonitoring ekspresi gen-gen stres dengan merancang primer multiplex Real Time-PCR pada gen housekeeping 18S Ribosomal RNA (18S rRNA), βeta Actin (βaktin) dan Elongation Factor 1-Alpha (EF1α), guna analisis ekspresi gen yang diuji pada organ hati, ginjal, dan usus ikan sidat. Isolasi RNA dilakukan pada ikan sidat menggunakan organ hati, ginjal dan usus. Sekuen spesifik ketiga gen housekeeping dirancang untuk RT-PCR pada laman primer3. Primer multiplex dianalisis menggunakan software FastPCR untuk mengamplifikasi gen housekeeping secara in silico. Studi pustaka seleksi gen housekeeping dilakukan dengan 3 gen kandidat housekeeping yaitu 18S rRNA, βaktin, dan EF1α untuk mendapatkan gen yang terekspresikan secara stabil. Hasil analisis primer multiplex secara in silico menunjukkan ketiga primer teramplifikasi dan menghasilkan amplikon berukuran <150 bp pada primer 18S rRNA & EF1α, dan 500 bp pada primer βaktin. Nilai deltaG berkisar antara 0 dan -2 kkal/mol. Nilai PCR efficiency ketiga primer >70 %. Studi pustaka seleksi gen housekeeping yang berpotensi memiliki stabilitas tinggi adalah gen 18S rRNA. Berdasarkan uji primer multiplex secara in silico ketiga gen dapat digunakan dalam analisis RT-PCR secara in vitro penelitian selanjutnya. Indonesia is one of the countries as an exporter of eel (Anguilla bicolor), the product is produced through the cultivation of seeds (glass eel) caught in river estuaries. Eel spawning is difficult to do, it can be caused by environmental conditions that interfere with the fish's metabolic system. Based on this, the purpose of this study was to monitor the survival of the eels by monitoring the expression of stress genes by designing primers. multiplex Real Time-PCR on genes housekeeping 18S Ribosomal RNA (18S rRNA), βeta Actin (βactin) and Elongation Factor 1-Alpha (EF1α), for analysis of gene expression tested in liver, kidney and intestines of eel. RNA isolation was carried out on eel using the liver, kidneys and intestines. The specific sequences of the three genes were housekeeping designed for RT-PCR on the primary page3. Primary multiplex analyzed using FastPCRsoftware to amplify genes housekeeping in silico.A literature study on gene selection was housekeeping carried out with 3candidate genes, housekeeping namely 18S rRNA, βactin, and EF1α to obtain genes that were expressed stably. The results of primer analysis multiplex by in silico showed that the three primers were amplified and produced amplicons of <150 bp in size 18S rRNA & EF1α primers, and 500 bp on βactin primers. The deltaG values range between 0 and -2 kcal / mol. The PCR efficiency value of the three primers is> 70%. The literature study on housekeeping gene selection that has the potential for high stability is the 18S rRNA gene. Based on the multiplex primer test, the in silico three genes can be used inRT-PCR analysis in vitro in future studies.

Item Type:	Thesis (S1)
Uncontrolled Keywords:	Anguilla bicolor, Gen Housekeeping, Primer Multiplex, Real Time-PCR
Subjects:	L Education > L Education (General) Q Science > QH Natural history > QH301 Biology S Agriculture > SH Aquaculture. Fisheries. Angling
Divisions:	Fakultas Pendidikan Matematika dan Ilmu Pengetahuan Alam > Jurusan Pendidikan Biologi > Program Studi Biologi (non kependidikan)
Depositing User:	Yusi Yustami
Date Deposited:	06 Jan 2021 03:39
Last Modified:	06 Jan 2021 03:39
URI:	http://repository.upi.edu/id/eprint/58011

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