PENENTUAN KONDISI OPTIMUM UNTUK ISOLASI PROTOPLAS DAUN Dendrobium Thong Chai Golden

• Main Author: Mumtahana, Aulia (Author)
• Format: Book
• Published: 2013-08-14.
• Subjects: Thesis; NonPeerReviewed
• Online Access: Link Metadata
• Notes: http://repository.upi.edu/152/1/S_BIO_0800601_TITLE.pdf; http://repository.upi.edu/152/2/S_BIO_0800601_ABSTRACT.pdf; http://repository.upi.edu/152/3/S_BIO_0800601_TABLE%20OF%20CONTENT.pdf; http://repository.upi.edu/152/4/S_BIO_0800601_CHAPTER1.pdf; http://repository.upi.edu/152/5/S_BIO_0800601_CHAPTER2.pdf; http://repository.upi.edu/152/6/S_BIO_0800601_CHAPTER3.pdf; http://repository.upi.edu/152/7/S_BIO_0800601_CHAPTER4.pdf; http://repository.upi.edu/152/8/S_BIO_0800601_CHAPTER5.pdf; http://repository.upi.edu/152/9/S_BIO_0800601_BIBLIOGRAPHY.pdf; http://repository.upi.edu/152/10/S_BIO_0800601_APPENDIX.pdf
• Keywords: Program Studi Biologi (non kependidikan)

Protoplas merupakan sumber material terbaik bagi pengembangan hibridisasi somatik pada anggrek. Penelitian ini bertujuan untuk memperoleh kondisi optimum untuk isolasi protoplas dari daun Dendrobium Thong Chai Golden dengan menentukan konsentrasi manitol sebagai osmotikum, kombinasi larutan enzim, dan lamanya inkubasi daun dalam larutan enzim. Penentuan konsentrasi manitol dilakukan menggunakan metode konvensional dengan menentukan konsentrasi yang menyebabkan kondisi incipient plasmolysis. Penelitian tentang pengaruh larutan enzim dan lamanya inkubasi daun dalam larutan enzim terhadap jumlah protoplas yang dihasilkan dilakukan berdasakan rancangan acak lengkap faktorial yang terdiri dari dua faktor. Faktor pertama adalah kombinasi enzim yang terdiri dari (1,5% selulase + 0,2% pektinase), E2 (1% selulase + 1% pektinase), dan E3 (1% selulase + 0,5% pektinase). Faktor kedua adalah lamanya lamanya inkubasi daun dalam larutan enzim yang terdiri dari 3, 5, 16, dan 21 jam. Eksplan adalah daun dari tanaman yang berumur satu tahun yang ditanam secara in vitro. Protoplas dikultur dalam medium ½ MS yang ditambahkan dengan kombinasi NAA dan BAP dan diamati setiap harinya selama dua minggu. Hasil menunjukan bahwa konsentrasi manitol terbaik sebagai osmotik adalah 0,5 M. Tidak ada interaksi pengaruh larutan enzim dan waktu inkubasi. Jumlah protoplas paling banyak (86,5 ± 22,7 x 104 protoplas/g berat basah) dihasilkan dari kombinasi enzim E1 dan inkubasi 16 jam. Duncan multiple range tests menunjukan tidak ada perbedaan signifikan dalam jumlah protoplas ketika daun diinkubasi selama 16 jam pada semua kombinasi enzim. Setelah dua minggu masa kultivasi dalam medium ½ MS yang ditambahkan seluruh kombinasi NAA dan BAP, plating efficiency kultur protoplas tidak dapat ditentukan. Kata Kunci : protoplas daun, Dendrobium Thong Chai Golden, osmotikum, kombinasi enzim,dan waktu inkubasi. Protoplast is the best source of material for improvement of somatic hybridization on orchids. This study was aimed to obtain an optimum condition for protoplast isolation of Dendrobium Thong Chai Golden leave protoplast by determining mannitol concentration as osmotic agent, combination of enzyme solution, and duration of leaves incubation in enzyme solution. Determination of mannitol concentration was conducted using conventional method by determining a concentration causing incipient plasmolysis. The study of the effect of enzyme solution and duration of leave incubation in enzyme solution on the number of released protoplasts was done according to factorial complete random design consisted of two factors. The first factor was enzyme combination consisted of E1 (1,5% cellulase + 0,2% pectinase), E2 (1% cellulase + 1% pectinase), and E3 (1% cellulase + 0,5% pectinase). The second factor was duration of leave incubation in enzyme solution consisted of 3, 5, 16 and 21 hours. The explan used was a one year old in vitro leaves. The protoplasts were cultured in ½ MS medium that was supplemented with several combination of NAA and BAP and observed every days for two weeks. The result showed that the best mannitol concentration as osmotic agent was 0,5 M. There were no interaction effect of enzyme solution and duration of incubation. The highest protoplast number (86,5 ± 22,7 x 104 protoplasts/ fresh weight) was yield by enzyme combination E1 and 16 hours incubation. Duncan multiple range tests showed No significant differences in protoplast number when the leaves were incubated for 16 hours in all enzyme combinations. After two weeks cultivation in ½ MS medium that was supplemented by all combination of NAA and BAP, plating efficiency of protoplast culture could not determined. Keywords : leave protoplast, Dendrobium Thong Chai Golden, osmotic agent, enzyme combination, and duration of incubation.