CRISPR/Cas9-mediated genome editing in rice cv. IPB3S results in a semi-dwarf phenotypic mutant

Nurhayati, Ardie SW, Santoso TJ, Sudarsono. 2021. CRISPR/Cas9-mediated genome editing in rice cv. IPB3S results in a semi-dwarf phenotypic mutant. Biodiversitas 22: 3792-3800. IPB3S is Indonesian lowland rice and high-yielding cultivar. However, plant height posture makes it prone to lodging which could reduce the yield. This study aimed to edit the GA20Ox2 gene by introducing CRISPR/Cas9 GA20Ox2 construct into IPB3S and developing the semidwarf rice mutants. Immature embryo explants of IPB3S were used for the transformation process mediated by EHA105 strain of Agrobacterium tumefaciens carrying pC1300-Cas9/GA20Ox2 and by changing the regeneration medium composition. PCR analysis showed that rice cv. IPB3S genetic transformation gained putative mutant T0 lines carrying the hpt gene (growth efficiency was 47.9%, while transformation efficiency was 19.3%). Using the developed regeneration medium, we have obtained 24 putative rice cv. IPB3S T0 mutant lines carrying hpt. The best medium for regenerating IPB3S was the A medium (regeneration efficiency 73.3%). IPB 8 and IPB 14 were potential to evaluate in the next generation. The shortest plant height for the T1 generation was observed in the IPB 8-3 mutant.

ANALISIS LINTAS PERTUMBUHAN DAN PRODUKSI TERHADAP PROTEIN KACANG TUNGGAK (Vigna unguiculata L) GENERASI M2

Cowpea (V. unguiculata L) can be developed as a source of vegetable protein because it contains relatively high protein and high lysine. The protein content in seeds is an accumulation of character components that play a role in protein formation, whose relationship can be predicted using cross correlation and analysis coefficients. The study was carried out in the experimental garden of Pasir Kuda PKHT of IPB in February - May 2018. Protein analysis is carriedout in the Testing Laboratory of the Department of Agronomy and Horticulture, Faculty of Agriculture, IPB in June - July 2018. The study was conducted using 30 putative mutant test genotypes of M2 generation result from the mutation of gamma ray irradiation and KM4 genotype as a comparison which was repeated three times. The protein content of the seeds was analyzed using the Kjeldahl method and then analyzed the variance, correlation coefficient and path. The results showed that M2 generation putative mutants showed diversity for the character of protein content in cowpea seeds. The protein content in cowpea seeds can be predicted through the character of plant height and number of branches.

ANALISIS KERAGAMAN DAN KEKERABATAN KACANG TUNGGAK (Vigna unguiculata L) GENERASI M2

Cowpea (V. unguiculata L) has great potential as a nutritious food as a substitute for soybeans because it contains sufficient protein and low fat content. The diversity of cowpea is low so that need to increase diversity through the mutation induction of gamma ray irradiation. The study was carried out in the experimental garden of Pasir Kuda PKHT of IPB in February - May 2018 using a design of augmented in the Complete Group Design in a Randomized. The study was carried out using 90 putative mutant genotypes of M2 generation as the test genotype and KM4 genotype as a comparison which was repeated 10 times. The results showed that the M2 generation putative mutants showed diversity in the qualitative and quantitative characters of cowpea. High genetic diversity is shown in the character of plant height, harvest period, number of seeds / pods and weight of cowpea seeds / plants and high broad mean heritability values obtained on the character of stem length, flowering age, number of seeds / pods and weight of beans / plant nuts arrears. The result of kinship analysis showed thirteen different putative mutant genotypes with KM4 genotypes, namely T6599P, T8028P, T7525P, T7551P, T7520P, T6574P, T6533P, T7058P, T6577P, T6591P, T7062P, T7069P and T6561.

Pengaruh Mutasi Fisik Iradiasi Sinar Gamma terhadap Keragaman Genetik dan Penampilan Coleus blumei

<p>ABSTRACT<br />Mutation breeding such as gamma ray irradiation is one of strategy to increase genetic variability. The aim of this research was to indentify genetic variability, performance changes and to obtain putative mutant of Coleus blumei purple/green through gamma ray irradiation. The experiment design used was Randomized Complete Block with single factor and three replications. The gamma ray irradiation was given to shoot cuttings of C. blumei by fractionated irradiation dose: 0 Gy (control), 20+20 Gy, 22.5+22.5 Gy, 25+25 Gy and 27.5+27.5 Gy. The irradiated shoot cuttings were planted in field until MV3 generation. The result of this research showed that gamma ray irradiation on C.blumei purple/green produced the high genetic variability on number of leaves and number of branches i.e. 58.48% and 74.02% by 25+25 Gy dose and number of branches by 20+20 Gy and 22.5+22.5 Gy dose i.e. 53.47% and 68.97% respectively. Physically induced mutation by gamma ray irradiation produced 5 putative mutants respectively on colour and pattern of leaf changes in the following plants: 20+20.5, 20+20.7, 22.5+22.5.8, 25+25.5 and 25+25.8.<br />Keywords: fractionated irradiation, mutagen, ornamental plant, putative mutan, shoot cutting</p><p>ABSTRAK<br />Pemuliaan mutasi dengan iradiasi sinar gamma merupakan salah satu cara dalam meningkatkan keragaman genetik tanaman. Tujuan penelitian ini untuk mengidentifikasi keragaman genetik, perubahan penampilan dan mendapatkan mutan putatif pada tanaman Coleus blumei ungu/hijau melalui iradiasi sinar gamma. Penelitian menggunakan rancangan kelompok lengkap teracak (RKLT) faktor tunggal dengan 3 ulangan. Iradiasi sinar gamma diberikan terhadap stek pucuk C. blumei ungu/hijau dengan dosis terbagi yaitu: 0 Gy (kontrol), 20+20 Gy, 22.5+22.5 Gy, 25+25 Gy dan 27.5+27.5 Gy. Semua tanaman hasil iradiasi ditanam di lapangan sampai generasi MV3. Hasil penelitian menunjukkan bahwa pemberian iradiasi sinar gamma pada C. blumei ungu/hijau menghasilkan keragaman genetik yang cukup tinggi pada karakter jumlah daun dan jumlah cabang dengan nilai KKG masing-masing 58.48% dan 74.02% pada dosis 25+25 Gy serta karakter jumlah cabang dengan nilai KKG 53.47% dan 68.97% masing-masing pada dosis 20+20 gy dan 22.5+22.5 Gy. Mutasi induksi fisik dengan iradiasi sinar gamma pada C. blumei ungu/hijau<br />menghasilkan 5 mutan putatif berdasarkan perubahan warna dan corak daun yaitu pada tanaman: 20+20.5, 20+20.7, 22.5+22.5.8, 25+25.5 dan 25+25.8.<br />Kata kunci: iradiasi terbagi, mutagen, mutan putatif, stek pucuk, tanaman hias</p>

ANALYSIS OF GAMMA IRRADIATED FOURTH GENERATION MUTANT OF RODENT TUBER (TYPHONIUM FLAGELLIFORME LODD.) BASED ON MORPHOLOGY AND RAPD MARKERS

Rodent tuber (Typhonium flagelliforme Lodd.) is an herbal medicinal plant with anticancer activity. The genetic diversity of rodent tuber is low due to vegetative propagation. Somatic cell population of rodent tuber from Bogor had been irradiated with gamma ray to increase genetic diversity. There were 37 clones of first generation putative mutant (MV1) which had been analyzed based on morphological and RAPD markers. Out of those 37 MV1 clones, there were 17 clones which had undergone genetic mutation and had a diversified genetic profile. MV1 had been regenerated to fourth generation putative mutant clones (MV4). This research was aimed to analyze the mutation stability of MV4 based on morphological and RAPD markers. Clone 6-1-2 had the highest increase of the number of shoots and leaves than control and the other MV4 clones, with 4.7 and 19.7 shoots and leaves, respectively. Clone 6-1-1-6 obtained the highest increase in plant height than control and the other MV4 clones, i.e. 25.2 cm. Clone 6-9-5 had the weightiest fresh and dry weight, i.e. 41.67 gram and 12.01 gram respectively. RAPD molecular marker analysis of MV4 by using 15 primers had produced 64 polymorphic DNA bands out of 146 total bands. OPD-10 primer produced the highest number of polymorphic bands, i.e. 15 polymorphic bands out of 17 total bands with sizes 200-2000 bp. RAPD profile of MV4 had showed 5 main clusters at similiarity coefficient cut-off 0.91. Morphological characterization and RAPD analysis had proved the stability of genetic mutation of MV4.

Keragaan dan Keragaman Genetik Karakter Agronomi Galur Mutan Putatif Gandum Generasi M5

The cultivation of wheat in Indonesia especially at medium-low altitude faces problems such as high temperatures whicheffect negativelyon the yield. The aims of this research was to obtain informations on performance, genetic variability and heritability of agronomic characters of wheat putative mutant lines on M5 generation. Genotypes evaluated in the study consisted of 98 putative mutant lines of M5 generation and six varieties as checks, namely Dewata, Selayar, Oasis, Rabe, Kasifbey, and Basribey. The research was conducted at Research Station of IOCRI (Indonesian Ornamental Crops Research Institute), Cipanas. Genotypes were arranged in an augmented, randomized block design. Result of the experiment showed that high heritability estimates were observed for plant height (83.03%), tiller number (61.75%), days to flowering (85.41%), days to maturity (67.45%), spike length (70.25%), spikelet number (65.08%), unfilled grain number (71.49%), seed number per spike (75.16%), seed weight per spike (81.69%), and seed weight per plant (74.21%). The estimated genotypic coefficient of variation (GCV) were high for unfilled grain number(26.35%) and seed weight per plant (18.35%). Based on seed weight per plant, 30 best putative mutant lines were selected.<br />Keywords: Augmented design, heritability, Triticum aestivum

KERAGAAN GENERASI KETIGA (M3) KEDELAI HASIL IRADIASI SINAR GAMMA PADA KONDISI OPTIMUM DAN KONDISI CEKAMAN KEKERINGAN

The objective of this research was to obtain a genotype of selected putative mutant withgood agronomic traits and high yielding on the optimum condition and its adaptability todrought stress in M3 generation. The study result showed that there were differences in thegrowth and production of each population as a result of irradiation in optimum conditions anddrought stress conditions. The population as a result of irradiation at 150 Gy and 200 Gy had a highermean value for the plant height trait, the number of productive branches, the number of productivenodes and the number of productive pods compared to irradiation population of 50 Gy, 100 Gy andthe control population of Argomulyo at optimum conditions. The decrease in the growth anddevelopment happened to all the characters observed in the irradiation population in droughtconditions compared to when grown in optimum conditions. The decrease can be seen in thecharacters of plant height, the number of pods and seed weight per plant. The growth and developmentof the population irradiation of 150 Gy and 200 Gy were more influenced by drought conditionscompared to other irradiation populations. The selection in the optimum conditions produced 291high yielding genotypes and the selection in drought conditions generated 202 genotypes andadapted to drought environments.

Identifikasi Genetik Mutan Talinum paniculatum JACQ. (GAERTN.) Berdasarkan Marka RAPD

Javasom Talinum paniculatum Jacq. (Gaertn.) is one of popular vegetable plants with potential medicinal properties.The plant root is often used as a substitute for ginseng (Panax ginseng L.), and the leaves are used as a vegetableand as a substitute for purslane (Portulaca olaraeae L.). An effort to improve genetic quality of the plant was madethrough induced mutation with ethyl methane sulphonate (EMS). A variety of EMS dosages (0, 0.3%, 0.6%, 0.9%,1.2%, 1.5% dan 1.8%) were applied to javasom seeds to induce mutation for 24 hours at room temperature. Selectedfive putative mutant genotypes of the third generation after mutagen treatment (M3) was analyzed using Ramdomamplified polymorphic DNA (RAD) marker. Twenty RAPD primers generated 185 scorable bands with 78.92% ofthem were polymorphic. Size of the bands varied from 200bp to 2.5kbp. Clustering analysis was performed basedon RAPD profiles using the UPGMA method. The range of genetic distance among individual genotypes was fromfrom 0.31 to 0.58, while genetic variance was 0.29 + 0.17. The five genotypes were proof to be solid mutants.