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16s rRNA Identification of Pediococcus spp. from Broiler and Studies of Adherence Ability on Immobilized Mucus Damayanti, Ema; Yusiati, Lies Mira; Dinoto, Achmad
Indonesian Journal of Biotechnology Vol 17, No 2 (2012)
Publisher : Universitas Gadjah Mada

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The objectives of this research were to study taxonomical status of lactic acid bacteria (LAB) isolated from broiler and adherence ability on mucus in vitro. Molecular analysis was performed by analyzing 16S rRNA gene using universal primer. The adherence assay on mucus was carried out using microplate method with total plate count (TPC), absorbance (A550) and confirmed by scanning electron microscopy (SEM). The results of this studies revealed that three of LAB isolates have closed relation to Pediococcus acidilactici (99.9%) species.Three isolates of P. acidilactici have adherence ability on broiler mucus higher than that on porcine mucin with an adherence percentage of 55.5% versus 50.8% and absorbance A550 of 0.061 versus 0.051, respectively. The highest adherence ability showed by P. acidilactici R02 with adherence percentage was 59.3% and absorbance A550 = 0.068. Adherence on mucus were affected by the addition of 3 g/l of gastric juice and 0.3% (b/v) of bile salt. Adherence analysis using SEM also showed that the adherence on broiler mucus was higher than the adherence on porcine mucin. Altogether this adherence studies, suggest that three isolates of P. acidilactici LAB were capable of colonizing host intestinal mucus in vitro as important property to be promising probiotic bacteria for broiler.Key words : adherence, broiler, Pediococcus, mucus, 16S rRNA

Identification of Pediococcus Strains Isolated from Feces of Indonesian Infants With in vitro Capability to Consume Prebiotic Inulin and to Adhere on Mucus ., Widodo; Anindita, Nosa Septiana; Taufiq, Tiyas Tono; Wahyuningsih, Tutik Dwi
Indonesian Journal of Biotechnology Vol 17, No 2 (2012)
Publisher : Universitas Gadjah Mada

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The aim of this experiment was to identify isolates obtained from feces of Indonesian infants and to evaluate their capability as probiotics. Identification of isolates was carried out based on morphology, physiology and biochemical identifications, and molecular identification based on 16S rRNA sequence. Morphological and physiological identification was carried out based on Gram staining, shape, motility, spore formation and catalase production. Biochemical identifications based on production of CO2 and NH3 from glucose, the ability to grow on different temperature (10 and 45°C) and pH (4.4 and 9.6), and different salt concentration (6.5 and 18%). Probiotics capability of isolates was assayed on the ability to grow on low pH (pH 2.0), on different bile salts concentration (0.3; 0.5; 1.0 and 1.5%), the capacity to grow on media with inulin as the only carbon source, and in vitro adhesion ability on porcine mucin. Morphological, physiological and biochemical identification suggest that all of isolates belong to lactic acid bacteria. Further molecular identification of five isolates showedthat isolates AA, BE and BK were strains of Pediococcus acidilactici (similarity 99%), while isolate AP and AG were strains of Lactobacillus casei (similarity 99-100%). Probiotic assays showed that more than 80% of cells of Pediococcus acidilactici isolates AA, BE and BK were viable after grown on pH 2.0 for 90 min, and around 80% of cells from the same isolates were survived on media supplemented with bile salt 1.5% for 2 h. All of isolates had high adhesion capacity as seen by more than 75% of cells attached on pig gastric mucin. Investigationof isolates to grow on inulin showed Pediococcus acidilactici isolate BE was able to consume inulin as the only carbon source. It is concluded that Pediococcus acidilactici isolate BE was a candidate probiotics and subject to further in vivo evaluation using animal models to examine their beneficial health effects.Key word : Pediococcus acidilactici, Lactobacillus casei, human origin and probiotics.

Genetic Relatedness among Duku, Kokosan, and Pisitan in Indonesia Based on Random Amplified Polymorphic DNA Markers Hanum, Laila; Kasiamdari, Rina Sri; ., Santosa; ., Rugayah
Indonesian Journal of Biotechnology Vol 17, No 2 (2012)
Publisher : Universitas Gadjah Mada

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Genetic relatedness among duku, kokosan, and pisitan from Indonesia were investigated using random amplified polymorphic DNA (RAPD) markers. Eleven primers (OPA-01, OPA-02, OPA-10, OPB-07, OPB-11, OPB-12, OPB-15, OPT-16, OPU-14, OPU-19, and OPU-20) were used for amplification and yielded a total of 174 DNA bands, of which 167 were polymorphic. Primer OPA-10, OPB-11, OPB-12, OPB-15, and OPU-19 produced all of the polymorphic DNA bands. The size of the amplified DNA fragments ranged from 41-1546 bp. The dendrogram separated into two clusters at a genetic similarity coefficient of 0.76. The cluster 1 consisted of subclusters duku and several pisitan (pisitan OKI, pisitan Sleman, pisitan Hatu, pisitan Punggur, and pisitan Tanjung), and cluster 2 consisted of subclusters kokosan and pisitan. In the kokosan subclusters, including duku Drendan. Dendrogram supported the determination of taxonomic status of duku, kokosan, and pisitan as one species, namely Lansium domesticum Corr. and its divided into two groups, namely L. domesticum ’duku group’ and L. domesticum ’pisitan-kokosan group’. Thus, RAPD analysis was useful tool for determining the genetic variation and the genetics relatedness among duku, kokosan, and pisitan in Indonesia.Key words: duku, kokosan, pisitan/langsat, genetic relatedness, RAPD

Functional Analysis of OsKANADI1, A Florigen Hd3a Interacting Protein in Rice (Oryza sativa L.) Purwestri, Yekti Asih; Ogaki, Yuka; Tsuji, Hiroyuki; Shimamoto, Ko
Indonesian Journal of Biotechnology Vol 17, No 2 (2012)
Publisher : Universitas Gadjah Mada

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OsKANADI1 is considered as a florigen Hd3a interacting protein. To study the function of OsKANADI1, the expression pattern of OsKANADI1 was performed by semiquantitative RT-PCR with various wild-type tissues in the floral transition stage. The results demonstrated that OsKANADI1 was expressed in all organs of wild-type plants, but was highest in roots and leaves. We hypothesize that OsKANADI1 is a transcription factor in rice because it contains a GARP domain and posses a nuclear localization signal. To determine whether OsKANADI1 encodes a nuclear protein, full-length OsKANADI1 fused to GFP was introduced into onion epidermis cells by particle bombardment. The result revealed that OsKANADI1 was localized in the nucleus, suggesting that OsKANADI1 may be a transcription factor. Functional analysis was carried out using a reverse genetics approach to generate gain of function mutant (overexpression) and knockdown mutant (RNAi). The results showed that suppression of OsKANADI1 by RNAi displayed branching and increasing tiller number in several lines. This phenotype resembles to the Hd3a overexpressed plants indicating they possibly function in similar pathway.Key words : OsKANADI1, Transcription factor, Hd3a interacting protein, Rice

High Frequency Spontaneous Deletions within the IcaADBC Operon of Clinical Staphylococcus epidermidis Isolates. Nuryastuti, Titik; van der Mei, Henny C.; Busscher, Henk J.; Kuijer, Roel; Aman, Abu Tholib; P. Krom, Bastiaan
Indonesian Journal of Biotechnology Vol 17, No 2 (2012)
Publisher : Universitas Gadjah Mada

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Staphylococcus epidermidis has been shown to undergo a phase variation correlating with expression of the icaADBC operon which contributes to biofilm formation. Biofilm formation of Enterococcus faecalis is related to heterogeneity in electrophoretic mobility. Here the relationship between phase variants of clinical isolates of S. epidermidis, icaADBC presence and electrophoretic mobility distributions is investigated. Of 105 S. epidermidis clinical isolates, 5 showed phase variation on Congo Red agar plate. Biofilm forming capability of the blackcolonies and inability of the red colonies were confirmed using a microtiter plate assay and confocal laser scanning microscopy. Upon analysis of electrophoretic mobility distributions, the black colonies displayed heterogeneity at pH 2 which was absent in the red colonies of the same strain. Surprisingly, it was shown that in all red colonies had lost the icaADBC genes. Determination of gene copy number using Real Time PCR targeting icaA showed reduction of gene copy within a culture with phase variation. In conclusion, using three fundamentally different approaches phase variation of the five clinical isolates was observed. Variants appeared through loss of icaA and icaC gens. To our knowledge this is the first report indicating S. epidermidis strains irreversible switching from biofilm + to biofilm – phenotype by deletion of ica genes. Key words: deletion, ica genes, Staphylococcus epidermidis, IcaADBC operon

Paternity Analysis of Tea (Camellia sinensis L. Kuntz) Hybrids Using Isozyme Marker Setyorini, Titin; ., Taryono; ., Suyadi; Indrioko, Sapto
Indonesian Journal of Biotechnology Vol 17, No 2 (2012)
Publisher : Universitas Gadjah Mada

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Tea plant has been categorized as self-incompatible crop. This is the reason behind the high genetic diversity. Natural pollination is possible to occur and the male parent is usually unknown, therefore, there is a need of method to identify male parent of hybrids through paternity analysis. Isozyme markers have been successfully used for paternity analysis due to their co-dominant polymorphism. This research aimed to predict male parents of hybrids by figuring out the mating system through isozyme banding patterns. In this experiment, seven enzyme systems were evaluated, of which only two of the enzyme systems i.e. esterase and shikimate dehydrogenase showing clear band pattern of Est-1, Est-2, and Shd-1 loci. The mating system of tea could be categorized as a mixed mating model, with high estimated out-crossing rate of 98.6 %. The pollen contributors were not always originated from the vicinity of the female parents.Key words: isozyme markers, paternity analysis, tea

Genetic Variation Analysis of Mold (Magnaporthe oryzae B.Couch) Using Random Amplified Polymorphic DNA Pramono, Ajeng Kusumaningtyas; Daryono, Budi Setiadi
Indonesian Journal of Biotechnology Vol 17, No 2 (2012)
Publisher : Universitas Gadjah Mada

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Magnaporthe oryzae B.Couch is a host-specific fungi, certain strain only infect certain host plant species. Genetic variety among M. oryzae isolates was explained by dendogram which was constructed using similarity data of Random Amplified Polymorphic DNA (RAPD). Dendogram construction was achieved by computer software, Numerical Taxonomy System (NTSYS). The aim of the research were to study the genetic variation among M. Oryzae using RAPD and to construct a dendogram of genetic similarities among the ten isolates from green foxtail (Setaria viridis L.), finger millet (Eleusine coracana L.) and rice (Oryza sativa L.).RAPD was performed in 30 cycles using 5 primers (OPA-02, OPA-03, OPA-04, OPA-05, OPA-07). Polymorphism data was used to constructed dendogram using Dice index and Unweighted Pair Group Method with Arithmetic Mean (UPGMA) in NTSYS software. There were 68 polymorphism fragments from 74 amplified fragments.Three clusters were formed in the dendrogram, based on host pathotype: foxtail millet type, finger millet type and rice type. There were two subclusters in foxtail millet type based on mating type, MAT1-1 dan MAT1-2. Thus, RAPD could be used as a method for genetic variation analysis of Magnaporthe oryzae to show host-specific specificity.Key words: Magnaporthe oryzae, RAPD, mating type

Molecular Marker Confirmation for Member of Anopheles barbirostris Van Der Wulp 1884 in Different Localities Satoto, Tri Baskoro Tunggul
Indonesian Journal of Biotechnology Vol 17, No 2 (2012)
Publisher : Universitas Gadjah Mada

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Vector and non-vector forms of Anopheles barbirostris have been recognized in Indonesia. However, because of their similarity in morphology, they were considered to be a single species. This information has led to the hypothesis that Anopheles barbirostris is a complex of species, which are morphologically indistinguishable from each other by ordinary methods. Objectives of the research was to identify the member of Anopheles barbirostris by PCR Assay. Samples were taken from two localities in Java, two in Sulawesi, two in Flores Indonesia, one from Thailand, one from China. The study was to develop a PCR-based technique of rDNA ITS2 region. Results showed that there are at least four species within the Anopheles barbirostris population studied, namely Anopheles barbirostris species DW, DX, DY and DZ. The length of the sequence amplified for species W, species X, species Y, and species Z were 339bps, 247bps, 165bps. and 157bps, respectively. Verification of the method was carried out with 270 mosquitoes from eight different field-collection sites using various sampling methods. Samples collected from Singaraja-Flores were identified as species W and X. All specimens collected from human bite outdoors were identified as species X; this species showed to be predominant among indoor light trap, indoor human bite and indoor resting collections Samples from Reo-Flores were identified as species W and X. All specimens from Manado and Palopo in Sulawesiwere identified as species Z. Similarly only species Y was found in samples from Thailand, while specimens from Salaman and Jambu in Java were identified as species W or species X. These species-specific molecular markers for the Anopheles barbirostris, complex appear to be reliable over a wide geographical area. However, larger number of samples is still needed from throughout the range of this species.Key words: Anopheles barbirostris, ITS2, PCR, Specific primer diagnostic

Identification of Pediococcus Strains Isolated from Feces of Indonesian Infants With in vitro Capability to Consume Prebiotic Inulin and to Adhere on Mucus W. Widodo; Nosa Septiana Anindita; Tiyas Tono Taufiq; Tutik Dwi Wahyuningsih
Indonesian Journal of Biotechnology Vol 17, No 2 (2012)
Publisher : Universitas Gadjah Mada

The aim of this experiment was to identify isolates obtained from feces of Indonesian infants and to evaluate their capability as probiotics. Identification of isolates was carried out based on morphology, physiology and biochemical identifications, and molecular identification based on 16S rRNA sequence. Morphological and physiological identification was carried out based on Gram staining, shape, motility, spore formation and catalase production. Biochemical identifications based on production of CO2 and NH3 from glucose, the ability to grow on different temperature (10 and 45°C) and pH (4.4 and 9.6), and different salt concentration (6.5 and 18%). Probiotics capability of isolates was assayed on the ability to grow on low pH (pH 2.0), on different bile salts concentration (0.3; 0.5; 1.0 and 1.5%), the capacity to grow on media with inulin as the only carbon source, and in vitro adhesion ability on porcine mucin. Morphological, physiological and biochemical identification suggest that all of isolates belong to lactic acid bacteria. Further molecular identification of five isolates showedthat isolates AA, BE and BK were strains of Pediococcus acidilactici (similarity 99%), while isolate AP and AG were strains of Lactobacillus casei (similarity 99-100%). Probiotic assays showed that more than 80% of cells of Pediococcus acidilactici isolates AA, BE and BK were viable after grown on pH 2.0 for 90 min, and around 80% of cells from the same isolates were survived on media supplemented with bile salt 1.5% for 2 h. All of isolates had high adhesion capacity as seen by more than 75% of cells attached on pig gastric mucin. Investigationof isolates to grow on inulin showed Pediococcus acidilactici isolate BE was able to consume inulin as the only carbon source. It is concluded that Pediococcus acidilactici isolate BE was a candidate probiotics and subject to further in vivo evaluation using animal models to examine their beneficial health effects.Key word : Pediococcus acidilactici, Lactobacillus casei, human origin and probiotics.

Genetic Relatedness among Duku, Kokosan, and Pisitan in Indonesia Based on Random Amplified Polymorphic DNA Markers Laila Hanum; Rina Sri Kasiamdari; S. Santosa; R. Rugayah
Indonesian Journal of Biotechnology Vol 17, No 2 (2012)
Publisher : Universitas Gadjah Mada

Genetic relatedness among duku, kokosan, and pisitan from Indonesia were investigated using random amplified polymorphic DNA (RAPD) markers. Eleven primers (OPA-01, OPA-02, OPA-10, OPB-07, OPB-11, OPB-12, OPB-15, OPT-16, OPU-14, OPU-19, and OPU-20) were used for amplification and yielded a total of 174 DNA bands, of which 167 were polymorphic. Primer OPA-10, OPB-11, OPB-12, OPB-15, and OPU-19 produced all of the polymorphic DNA bands. The size of the amplified DNA fragments ranged from 41-1546 bp. The dendrogram separated into two clusters at a genetic similarity coefficient of 0.76. The cluster 1 consisted of subclusters duku and several pisitan (pisitan OKI, pisitan Sleman, pisitan Hatu, pisitan Punggur, and pisitan Tanjung), and cluster 2 consisted of subclusters kokosan and pisitan. In the kokosan subclusters, including duku Drendan. Dendrogram supported the determination of taxonomic status of duku, kokosan, and pisitan as one species, namely Lansium domesticum Corr. and its divided into two groups, namely L. domesticum ’duku group’ and L. domesticum ’pisitan-kokosan group’. Thus, RAPD analysis was useful tool for determining the genetic variation and the genetics relatedness among duku, kokosan, and pisitan in Indonesia.Key words: duku, kokosan, pisitan/langsat, genetic relatedness, RAPD

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Contact Name
-

Contact Email
-

Phone
-

Journal Mail Official
-

Editorial Address
-

Location
Kab. sleman,

Daerah istimewa yogyakarta

INDONESIA