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LEMBARHASIL PENII,AIAN SHAWAT SEBIDANG ATAA PEER REWEIY
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Molecular characterization of dunaliella salina and chlorella vulgaris fusant using 18SrDNA gene (Article)
,
Genetics Laboratory, Biology Department, Faculty of Mathematics and Natural Sciences, Diponegoro University, Jl.Prof. Soedarto, SH Tembalang, Semarang, 50275, IndonesiaMarine laboratory. Faculty of Faculty of Fisheries and Marine Science, Diponegoro University, Jl. Prof. Soedarto, SH
Tembalang, Semarang, 50275, Indonesia
AbstractProtoplast fusion was found to be an efficient method in improving carotenoid production from fusant ofcarotenogenic microalgae D. salina and C. vulgaris . Molecular characterization is needed for identifying thedominant parental genome in the fusant using ribosomal DNA sequences. The research was carried out by analyzingthe gene encodes for 18S rDNA of fusant and determining relationship of fusant with D. salina and C. vulgaris species from GenBank. Quantitative analysis showed that C. vulgaris was not remarkably dominant in fusant with 84 % similarity compare to D. salina with 82 % similarity. The result indicated that the fusant gained
both character from their parents. © 2016 Penerbit UTM Press. All rights reserved.
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Author keywords18SrDNA Chlorellla Dunaliella Protoplast fusion
Lee, Y.K., Tan, H.M. Interphylum Protoplast Fusion and Genetic Recombination of the Algae Porphyridium cruentum andDunaliella spp (1988) Journal of General Microbiology, 134, pp. 635-641. .
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Lu, Y., Kong, R., Hu, L.
(2012) World Journal of Microbiology and Biotechnology, 28 (4), pp. 1827-1830. . doi: 10.1007/s11274-011-0963-4
Santiago, C.M. Protoplast Fusion A New Tecnique for Genetic Manipulation and Breeding of Industrial Microorganisme. I.C (1982) Biotech, 5, pp. 435-440.
Pulz, O., Gross, W.
(2004) Applied Microbiology and Biotechnology, 65 (6), pp. 635-648. . doi: 10.1007/s00253-004-1647-x
Borowitzka, M.A., Borowitzka, L.J. Dunaliella (1988) Micro-Algal Biotechnology. . Borowitzka, M. A. (ed.), Cambridge: Cambridge University Press
Kusumaningrum, H.P., Zainuri, M. Optimization and Stability of Total Pigments Production of Fusan from Protoplast Fusion of MicroalgaeDunaliella and Chlorella in vivo. Attempts on Production of Sustainable Aquaculture Natural Food (2014) International Journal of Marine and Aquatic Resource Conservation and Co-Existance, 1 (1), pp. 1-5. .
Guedes, A.C., Malcata, F.X. Nutritional Value and Uses of Microalgae in Aquaculture (2012) Aquaculture. . Muchlisin, Z. A, Rijeka: InTech
Guedes, A.C., Amaro, H.M., Malcata, F.X.
(Open Access) (2011) Marine Drugs, 9 (4), pp. 625-644. .
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Panaiotov, S., Evstatieva, Y., Ilieva, S., Levterova, V., Brankova, N., Nikolova, D., Ivanova, A., (...), Atev, A.
(2009) Biotechnology and Biotechnological Equipment, 23, pp. 892-895. doi: 10.1080/13102818.2009.10818566
Zainuri, M., Kusdiyantini, E., Widjanarko, Soedarsono, J., Yuwono, T. Preliminary Study on the Use of Yeast Phaffia rhodozyma as Pigment Source on The Growth of Tiger Shrimp(Penaeus monodon Fabricius) (2003) Indonesian Journal of Marine Sciences, 8 (1), pp. 47-52.
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Kusumaningrum, H.P.; Genetics Laboratory, Biology Department, Faculty of Mathematics and Natural Sciences,Diponegoro University, Jl. Prof. Soedarto, SH Tembalang, Semarang, Indonesia; email: © Copyright 2016 Elsevier B.V., All rights reserved.
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78:4–2 (2016) 61–68 |www.jurnalteknologi.utm.my | eISSN 2180–3722 |
Jurnal
Teknologi
Full Paper
MOLECULAR CHARACTERIZATION OF
Dunaliella salina AND Chlorella vulgaris
FUSANT USING 18SrDNA GENE
Hermin Pancasakti Kusumaningruma*, Muhammad Zainurib
aGenetics Laboratory, Biology Department, Faculty of
Mathematics and Natural Sciences, Diponegoro University, Jl. Prof.
Soedarto, SH Tembalang, Semarang – 50275, Indonesia bMarine laboratory. Faculty of Faculty of Fisheries and Marine
Science, Diponegoro University, Jl. Prof. Soedarto, SH Tembalang,
Semarang – 50275, Indonesia
Article history
Received
5 November 2015
Received in revised form
11 February 2016
Accepted
15 February 2016
*Corresponding author
Graphical abstract
Abstract
Protoplast fusion was found to be an efficient method in improving carotenoid production
from fusant of carotenogenic microalgae D. salina and C. vulgaris. Molecular
characterization is needed for identifying the dominant parental genome in the fusant
using ribosomal DNA sequences. The research was carried out by analyzing the gene
encodes for 18S rDNA of fusant and determining relationship of fusant with D. salina and C.
vulgaris species from GenBank. Quantitative analysis showed that C. vulgaris was not
remarkably dominant in fusant with 84 % similarity compare to D. salina with 82 % similarity.
The result indicated that the fusant gained both character from their parents.
Keywords: Chlorellla, Dunaliella, protoplast fusion, 18SrDNA
© 2016 Penerbit UTM Press. All rights reserved
1.0 INTRODUCTION
Recent decades has shown remarkable developing of
the biotechnology of microalgae. Valuable product
for food and other applications will extend into
broader area. Production of genetically improved
strain by hybridization and somatic fusion on algae
have been reported in algae. Protoplast fusion was
done on intraspesific species green microalgae
Chlamydomonas reinhardii (P. A. Dang, 1888) [1]. The
protoplast fusion between two different phyla which
sexually incompatible also has been conducted
between the red alga Porphyridium cruentum (S.F.
Gray) Nägeli, 1849) with D. bardawil (Ben-Amotz &
Avron, 1982) or D. salina (Dunal) Teodoresco 1905 [2].
D. salina is the source of β-carotene and glycerol [3, 4].
Chlorella is widely used as a healthy food and feed
supplement, as well as in the pharmaceutical and
cosmetics industries [5]. Improvement of valuable
metabolites from Chlorella and Dunaliella microalgae
was done using biotechnological methods that
allowed somatic hybridization by protoplast fusion [6].
This technique is required because crosses between
two strains will not occur naturally and a diploid is
desired. This intergeneric fusion also enables nuclear
and cytoplasmic genomes to be combined, fully or
partially, at the intergeneric levels. Carotenogenic
microalgae which live in different environmental
salinity, namely Chlorella and Dunaliella have
produced stable hybrids possessing combined
characteristics of the parents [6]. Their application as
natural supplement for aquaculture animal was
potential for synthetic feed substitution since it
contains proteins, carbohydrates, lipids and vitamins,
carotenoid as antioxidants, and trace elements [6–9].
Supplementation feeding on Penaeus monodon
postlarvae with microalgae exhibited significant effect
on growth, weight, survival related to microbial
diseases resistance and immune response in high and
low salinity [10–12]. In the present study, fusant
62 Hermin Pancasakti Kusumaningrum & Muhammad Zainuri / Jurnal Teknologi (Sciences & Engineering) 78:4–2 (2016) 61–68
between D. salina and C. vulgaris were obtained. The
purpose of this study was conducting molecular
characterization on fusant using 18SrDNA to asses
genetic exchange among parental strains during
fusant formation. The methodologies were conducted
by isolation and amplification of 18SrDNA region of
fusant, followed by analysis on variable and conserved
sequences that were contained in the 18SrDNAs of the
organisms. The comparison of rDNA sequences
between parental and progeny is a potencial tool for
deducing dominancy and combination of each
parental to improve their advantages in fusant. The
present study expands on the use of DNA technology
for the genotype within the fusant comparing to
parental.
2.0 EXPERIMENTAL
2.1 D. salina and C. vulgaris
D. salina and C. vulgaris microalgae were obtained
from Brackishwater Aquaculture Development Centre
(BBPBAP) on Jepara, Indonesia. They were held in
seawater tanks, recirculated and aerated, with the
temperature set at 25 °C to 28 °C and salinity at 30 ‰
to 32 ‰. The tanks were cleaned daily. The
microalgae were cultivated using sea water enriched
with Walne media. A twocomponent gel, it is easy to
modify the molecular structure of either of the two
components.
2.2 Microalgae Media
Walne media for microalgae growth and cultivation
consist of H3BO3 3.36 g L1, NaNO3 10 g L1, FeCl3 0.15 g
L1, MnCl2.4H2O 0.36 g L1, Na2EDTA 45 mg L1, NaH2PO4
20 g L1, trace metal solution 1 mL L1, and distilled
water. Trace metal solution was consist of H3BO3 2.86 g
·L1; MnCl2.4H2O 1.81 g L1; ZnSO4.7H2O 0.222 g L1;
NaMoO4.5H2O 0.39 g L1; CuSO4.5H2O 0.079 g L1;
Co(NO3)2.6H2O 0.0494 g L1; pH 6.8. The ingredients
were dissolved in 200 mL of distilled water. The solution
was boiled for 10 min while adjusting the pH to 7.6 with
HCl or NaOH, filtered and bring to 1 L. Sterilization was
done by autoclaving at 15 lb in2 (103 kPa and 120 oC).
The medium was using by adding 0.1 mL solution to
each 10 mL of seawater [13, 14].
2.3 DNA Extraction
The DNA was extracted from 3 L media of the fusant in
logarithmic phase of growth. The preparation of a
DNA genomic of fusants was carried out by
modification of CTAB methods [15, 16]. DNA was kept
on -20 °C or used directly for PCR.
2.4 Amplification of 18SrDNA Microalgae
The 18SrDNA fragment was amplified using specific
primers. Sequence of forward primer was 5’
GTAGTCATATGCTTGTCT3’, reverse primer was 5’
GCTGGCACCASACTTGCCCT3’ [17]. PCR was carried
out in mixture containing 50 ng of genomic DNA,a 2.5
L PCR buffer (KAPA), a 10 mM concentration of
deoxynucleoside triphosphate mix (KAPA), 2.5 pmol of
each primer, and 0.625 U (1 U=1/60 micro katal) of Taq
Extra Hotstart DNA Polymerase and ddH2O until
volume 25 L. PCR conditions were performed
according to the PCR protocol using the
manufacturer’s instructions and protocols. To amplify
the 18SrDNA, amplification reactions were performed
on a T-Gradient thermocycler (Biometra GmbH,
Gottingen, Germany). Aliquots (1 L) of the reaction
mixtures were analyzed by 0.8 % horizontal agarose gel
electrophoresis to confirm the presence of product.
The PCR products were purified using the Gel PCR
Clean-Up System (Applied Biosystems, Foster, CA).
Sequencing reactions were performed using a Dye
Deoxy Terminator Cycle Sequencing Ready Reaction
Kit (Applied Biosystems, Foster City, CA), and
sequencing fragments were analyzed on a ABI Prism
377 DNA Sequencer.
2.5 Sequencing and Phylogenetic Analysis
The 18S rDNA partial fragment gene sequences from
the fusant were searched against GenBank using
BLAST as illustrated in Table 1. The nucleotides were
aligned using the program ClustalX, respectively.
Sequences containing fewer than 400 nucleotides or in
excess of 1 000 nucleotides were removed, and
sequences not belonging to D. salina and C. vulgaris
microalgal species were discarded from this study. A
phylogenetic tree was constructed using the
neighborjoining (NJ) algorithm [18] using Kimura’s
twoparameter model of sequence evolution, as
implemented in the MEGA5 program package [19].
The bootstrap consensus tree inferred from 1 000
replicates [20] is taken to represent the evolutionary
history of the taxa analyzed. Branches corresponding
to partitions reproduced in less than 50 % bootstrap
replicates are collapsed. The percentage of replicate
trees in which the associated taxa clustered together
in the bootstrap test are shown next to the branches.
The evolutionary distances were computed using the
Jukes-Cantor method [21] and are in the units of the
number of base substitutions per site. Codon positions
included were 1st + 2nd + 3rd . All positions containing
gaps and missing data were eliminated.
63 Hermin Pancasakti Kusumaningrum & Muhammad Zainuri / Jurnal Teknologi (Sciences & Engineering) 78:4–2 (2016) 61–68
Table 1 Dunaliella and Chlorella species rDNA sequence from Genbank used in this study
Species Accs number Species Accs number
C. vulgaris AB080308.1 D. salina strain KU13 KF825552.1
C. vulgaris isolate UMT-M1 KJ561358.1 D. salina strain KU07 KF825551.1
C. vulgaris isolate KS-MA2 KJ561357.1 D. salina strain KU11 KF825550.1
C. vulgaris KF574391.1 D. salina strain KU07 KF825551.1
C. vulgaris cc CCAP 211/79 FR865683.1 D. salina isolate B32 HQ735296.1
C. vulgaris strain KTP2 KF746940.1 D. salina isolate I3 HQ735295.1
C. vulgaris strain nm27 JQ256478.1| D. salina strain B34 JF900404.1
C. vulgaris isolate YL-2 KP341004.1 D. salina strain B24 JF831044.1
C. vulgaris strain A1-65 KF661335.1 D. salina EF195157.1
C. vulgaris strain CCAP211/21A KJ756823.1 D. salina strain CCAP 19/12 KJ756842.1
C. vulgaris strain CCAP211/75 KJ756813.1 D. salina str BuriRam KU01 JN052202.1
C. vulgaris strain LC9 KF569735.1 D. salina strain I2 JF831045.1
C. vulgaris strain LC8 KF569734.1 D. salina strain CCAP 19/18 EF473745.1
C. vulgaris strain LF5 KF569724.1 D. salina strain SAG 19-3 EF473739.1
C. vulgaris strain AG-35_ZF1 AB699112.1 D. salina strain Dsge EF473731.1
C. vulgaris cc CCAP211/11P FR865658.1 D. salina strain KMMCC 1428 JQ315781.1
C. vulgaris strain CCAP211/110 FN298918.1 D. salina M84320.1
C. vulgaris strain CCAP211/109 FN298917.1 D. salina strain CCC HQ843776.1
C. vulgaris strain CCAP211/82 FM205855.1 D. salina strain JR102 EU589200.1
C. vulgaris strain CCAP211/81 FM205854.1 D. salina EU239363.1
C. vulgaris strain CCAP211/80 AM231734.1 D. salina AF506698.1
C. vulgaris strain CCAP 211/11F AY591515.1 D. salina M84320.1
C. vulgaris strain CCAP 211/63 FR865681.1 D. salina strain CCC HQ843776.1
C. vulgaris strain SAG 211-11b FM205832.1 D. salina strain JR102 EU589200.1
C. vulgaris strain KMMCC FC-15 HQ702287.1 D. salina EU239363.1
C. vulgaris strain KMMCC FC-12 HQ702286.1 D. salina AF506698.1
C. vulgaris strain KMMCC FC-12 HQ702286.1 D. salina M84320.1
C. vulgaris strain KMMCC C-111 GQ122346.1 D. salina strain CCC HQ843776.1
C. vulgaris strain KMMCCFC-16 HQ702294.1 D. salina strain JR102 EU589200.1
C. vulgaris strain KMMCCC-117 HQ702318.1 D. salina strain B33 JF831042.1
C. vulgaris strain KMMCCC-119 HQ702309.1 D. salina strain UTEX LB 200 DQ009779.1
C. vulgaris strain KMMCCFC-42 HQ702285.1 Dunaliella sp. BBPBAP KC875350
C. vulgaris strain KMMCCEC-5 HQ702321.1 D. salina isolate NIOT-28(ANCOST-17) KC470060.1
C. vulgaris strain KMMCCFC-33 HQ702295.1 D. salina strain CCAP 19/30 DQ447648.1
C. vulgaris cc KMMCC FC-1 GQ122369.1 C. vulgaris cc KMMCC C-27 GQ122334.1
C. vulgaris strain NIES-1269 AB488579.1 C. vulgaris var vulgaris strNIES-642 AB488577.1
C. vulgaris strain: PS-2670 AB488582.1 C. vulgaris var vulgaris strNIES-227 AB488575.1
Chlorella sp. WO10-1 FJ946886.1 C. vulgaris var vulgaris str NIES-641 AB488576.1
C. vulgaris strain KMMCCEC-10 HQ702292.1 C. vulgaris var vulgaris str NIES-686 AB488578.1
C. vulgaris strain KMMCCEC-3 HQ702293.1 C.vulgaris isolate D2 JX185298.1
C. vulgaris strain KMMCCFC-16 HQ702294.1 C. vulgaris strain IAM C-27 AJ242757.1
C. vulgaris strain KMMCCC-119 HQ702309.1 C. vulgaris strain KMMCCC-111 GQ122346.1
C. vulgaris strain KMMCCFC-42 HQ702285.1 C. vulgaris cc KMMCC C-88 GQ122340.1 Note: cc = culture collection; v= varian; accs = accession
3.0 RESULT AND DISCUSSION
3.1 Amplification of 18SrDNA fusant
The products of the 18SrDNA fusant amplification are
shown in Figure 1. Several annealing temperature were
applied due to anticipation of genetic diversity inside
fusant genome. Almost all temperature annealing
showed positive bands with 55.2 °C exhibited the best
result. Those data confirmed that primer design was
suitable to conserved region of fusant.
Figure 1 Amplification of 18SrDNA fusant with several
annealing temperature using gradient PCR. Lane 1:
molecular weight marker. Lanes 2 to 10: amplification with
annealing temperature 45 °C; 46.6 °C; 48.4 °C; 50.9 °C; 52.9
°C; 55.2 °C; 57.2 °C; 59.9 °C; 61.7 °C; and 63 °C, respectively
M 1 2 3 4 5 6 7 8 9 10 M
64 Hermin Pancasakti Kusumaningrum & Muhammad Zainuri / Jurnal Teknologi (Sciences & Engineering) 78:4–2 (2016) 61–68
Furthermore, the primer also can be used to amplify
fusant and also obtained similar band from parental
D. salina confirmed that primers are highly specific
to D. salina. On the other hand, by applying primer on
parental C. vulgaris single band was obtained but the
sequences were not as good as D. salina and fusant.
This result indicated that the primer was not in the
conserved region of C. vulgaris in amplifying specific
band. Implication of this result also showed the
divergence of sequences in C. vulgaris comparing
with D. salina. However, this result has not interfered
with the homology analysis of fusant sequences since
C. vulgaris showed conserved region among them.
Fusant 19 ATTGTACTCATTCCGATT-GCAGAACCCGAAGGGCTCCGCATCGTTATTTATTGTCACTACCTCCCTG 85
|||||||||||||| ||| ||| | | || || | | || ||||||||||||||||||||||| |
D.salina 469 ATTGTACTCATTCCAATTACCAG-A-CAAAAATGCCCGGTATTGTTATTTATTGTCACTACCTCCCCG 402
Fusant 86 TGTTAGGATTGGGTAATTTACGCGCCTGCTGCCTTCCTTAGATGTGGTAGCCGTTTCTCAGGCTCCCT 153
|||| |||||||||||||| ||||||||||||||||||| ||||||||||||||||||||||||||||
D.salina 401 TGTTGGGATTGGGTAATTTGCGCGCCTGCTGCCTTCCTTGGATGTGGTAGCCGTTTCTCAGGCTCCCT 334
Fusant 154 CTCCGGAATCGAACCCTAATTCTCCGTTACCCGTTAACGCCACGGTAGGCCAATACCCTACCGTCGAA 221
|||||||||||||||||||| |||||| |||||||| | ||| |||||||| || |||||| |||||
D.salina 333 CTCCGGAATCGAACCCTAATCCTCCGTCACCCGTTACCACCATGGTAGGCCTCTATCCTACCATCGAA 266
Fusant 222 AGCTGATAGGGCAGAAACTTGAATGAACCATCGT-GCCG-AA-GCA---CGATTCGCTTAGTTATTAT 283
|| |||||||||||||| ||||||||| ||||| || || || ||||||| |||||| |||
D.salina 265 AGTTGATAGGGCAGAAATTTGAATGAAACATCGCCGGCATAAAGCCGTGCGATTCGTGAAGTTATCAT 198
Fusant 284 GACTCACCA-G-G--G---ATT---G-CTGG--TTGTATCTAATAAATACACCTCTTGC-GAGGTTGG 337
|| || ||| | | | | | |||| || |||||||||||||| | ||| | || || ||
D.salina 197 GATTCGCCAAGAGTCGGGCAAGCCCGGCTGGCCTTTTATCTAATAAATACGTCCCTTCCAGAAGTCGG 130
Fusant 338 -----ACGCATGTATTAGCTCTAGAATTACTACGGTTATCCAAGTAGTAGGGGACTATCAAATAAACT 400
||||| |||||||||||||||||||||||||||||| |||| ||| || ||||||||||||
D.salina 131 GATTTACGCACGTATTAGCTCTAGAATTACTACGGTTATCCGAGTAA-AGGT-ACCATCAAATAAACT 66
Fusant 401 ATAACTGATATAATGAGCCATTCGCAGTTTCACCGTATAA-AGGCTTATACTTAG-ACATGCA 461
||||||||| ||||||||||||||||||||||| |||||| | |||||||||| |||||||
D.salina 65 ATAACTGATTTAATGAGCCATTCGCAGTTTCACAGTATAAGCAGTTTATACTTAGGACATGCA 3
Figure 2 Similarity among fusant and D. Salina
Fusant 1 CATGTCT-AGTAT-AGC--CTTTATACGGTG-AACTGCGAATGGCTCATTATATCAGTTATAGTTTAT 63
||||||| ||||| | | |||||||| ||| ||||||||||||||||||| ||||||||||||||||
C.vulgaris 53 CATGTCTAAGTATAAACTGCTTTATACTGTGAAACTGCGAATGGCTCATTAAATCAGTTATAGTTTAT 120
Fusant 64 TTGATAGTCCCCTACTACTTGGATAACCGTAGTAATTCTAGAGCTAATACATGCGT-----CCAACCT 131
||||| || ||||||||| ||||| ||||||||| |||||||||||||| ||||| || || |
C.vulgaris 121 TTGATGGT-ACCTACTACTCGGATACCCGTAGTAAATCTAGAGCTAATACGTGCGTAAATCCCGACTT 187
Fusant 132 C-GCAAGAGGTGTATTTATTAGATACAA--CC-A----GCAAT--CC--CT---GGTGAGTCATAATA 185
| | ||| | |||||||||||||| || || | || | || || ||||| |||| |||
C.vulgaris 172 CTGGAAGGGACGTATTTATTAGATAAAAGGCCGACCGGGCTCTGCCCGACTCGCGGTGAATCATGATA 255
Fusant 186 ACTAAGCGAATCG--T-G-CTT-CGGC-ACGATGGTTCATTCAAGTTTCTGCCCTATCAGCTTTCGAC 241
||| ||||||| | | ||| || | ||||| ||||||||| |||||||||||||| |||||||
C.vulgaris 256 ACTTCACGAATCGCATGGCCTTGCGCCGGCGATGTTTCATTCAAATTTCTGCCCTATCAACTTTCGAT 323
Fusant 242 GGTAGGGTATTGGCCTACCGTGGCGTTAACGGGTAACGGAGAATTAGGGTTCGATTCCGGAGAGGGAG 309
|||||| || |||||||| ||| | |||||||| |||||| ||||||||||||||||||||||||||
C.vulgaris 324 GGTAGGATAGAGGCCTACCATGGTGGTAACGGGTGACGGAGGATTAGGGTTCGATTCCGGAGAGGGAG 391
Fusant 270 CCTGAGAAACGGCTACCACATCTAAGGAAGGCAGCAGGCGCGTAAATTACCCAATCCTAACACAGGGA 377
|||||||||||||||||||||| ||||||||||||||||||| ||||||||||||||| |||||||||
C.vulgaris 352 CCTGAGAAACGGCTACCACATCCAAGGAAGGCAGCAGGCGCGCAAATTACCCAATCCTGACACAGGGA 459
Fusant 330 GGTAGTGACAATAAATAACGATGCGGAGCCCTT-CGGGTTCTGC-AATCGGAATGAGTACAATTTAAA 443
||||||||||||||||||| || | | ||| || | ||| ||| ||| |||||||||||||| ||||
C.vulgaris 412 GGTAGTGACAATAAATAACAATACTGGGCCTTTTCAGGT-CTGGTAATTGGAATGAGTACAATCTAAA 525
Fusant 390 CCCCTTAACGAGGATCCATTGGAGGGCAAGTCTGGTGCCAGCAG 487
|||||||||||||||| |||||||||||||||||||||||||||
C.vulgaris 472 CCCCTTAACGAGGATCAATTGGAGGGCAAGTCTGGTGCCAGCAG 570
Figure 3 Similarity among fusant and C. vulgaris
65 Hermin Pancasakti Kusumaningrum & Muhammad Zainuri / Jurnal Teknologi (Sciences & Engineering) 78:4–2 (2016) 61–68
3.2 Sequence Analysis of Fusant
Sequence analysis of 18S rDNA fragmen nucleic acid
in GenBank GenBank and European Bioinformatics
shows close relationship between fusant and D. salina
strain BBPBAP and C. vulgaris. Analysis of homology
between fusan and D. salina showed 82 % homology
(Figure 2). Analysis of homology between fusan and C.
vulgaris showed 84 % homology (Figure 3). Identities
founded 320 in 435 with gaps 6 %.
Homology analysis inside the 18SrDNA sequences of
fusant and parental identified 77 % similarity in the
region as illustrated in Figure 4. Both D. salina and C.
vulgaris exhibited conservation region between
sequences and the similarity among them was 94 % in
the 18SrDNA region which suggested that this
sequence was well conserved between species
of Dunaliella and Chlorella. Analysis and comparison
of sequences on fusant and parental detected 77 %
similarity with 33 SNPs and 24 deletion in the 18SrDNA
region (Figure 4). A number of base were inherited
between two parents into the fusant exhibiting by
several substitution. Combination of bases from two
parents into the fusant made differences from both
parent but C. vulgaris tend to inherited more dominant
comparing with D. salina.
C.vulgaris : C-TAAGTATAAACTGCTTTATACTGTGAAACTGCGAATGGCTCATTAAATCAGTTATAGTTTATTTGAT : 69
D.salina : CCTAAGTATAAACTGCTT-ATACTGTGAAACTGCGAATGGCTCATTAAATCAGTTATAGTTTATTTGAT : 70
Fusant : CATGTCTAGTATAGCCTTTATACGGTGAA-CTGCGAATGGCTCATTATATCAGTTATAGTTTATTTGAT : 70
C TaagTAtaAactgCTTtATACtGTGAAaCTGCGAATGGCTCATTAaATCAGTTATAGTTTATTTGAT 70
80 * 100 * 120 *
C.vulgaris : GGT-ACCTACTACTCGGATACCCGTAGTAAATCTAGAGCTAATACGTGCGTAAATCCCGACTTCTGGAA :139
D.salina : GGTACCTT--TACTCGGATAACCGTAGTAATTCTAGAGCTAATACGTGCGTAAATCCCGACTTCTGGAA :137
Fusant : AGTCCCCTACTACTTGGATAACCGTAGTAATTCTAGAGCTAATACATGCGT-----CCAACCTC-GCAA :135
gGT cCcTacTACTcGGATAaCCGTAGTAAtTCTAGAGCTAATACgTGCGTaaatcCCgACtTCtGgAA 69
140 * 160 * 180 * 200
C.vulgaris : GGGACGTATTTATTAGATAAAAGGCCGACCGGGCTCTGCCCGACTCGCGGTGAATCATGATAACTTCAC :208
D.salina : GGGACGTATTTATTAGATAAAAGGCCAGCCGGGCT-TGCCCGACTCTTGGCGAATCATGATAACTTCAC :209
Fusant : GAGGTGTATTTATTAGATA------CAACCAG--------CAATCCCTGGTGAGTCATAATAACTAAGC :196
GgGacGTATTTATTAGATAaaaggcCaaCCgGgct tgccCgActC tGGtGAaTCATgATAACTtcaC 68
* 220 * 240 * 260 *
C.vulgaris : GAATCGCATGGCCTTGCGCCGGCGATGTTTCATTCAAATTTCTGCCCTATCAACTTTCGATGGTAGGAT :278
D.salina : GAATCGCACGGCTTTATGCCGGCGATGTTTCATTCAAATTTCTGCCCTATCAACTTTCGATGGTAGGAT :279
Fusant : GAATCGTG----CTTCGGC--ACGATGGTTCATTCAAGTTTCTGCCCTATCAGCTTTCGACGGTAGGGT :260
GAATCGca ggccTT GCcggCGATGtTTCATTCAAaTTTCTGCCCTATCAaCTTTCGAtGGTAGGaT 67
280 * 300 * 320 * 340
C.vulgaris : AGAGGCCTACCATGGTGGTAACGGGTGACGGAGGATTAGGGTTCGATTCCGGAGAGGGAGCCTGAGAAA :348
D.salina : AGAGGCCTACCATGGTGGTAACGGGTGACGGAGGATTAGGGTTCGATTCCGGAGAGGGAGCCTGAGAAA :349
Fusant : ATTGGCCTACCGTGGCGTTAACGGGTAACGGAGAATTAGGGTTCGATTCCGGAGAGGGAGCCTGAGAAA :330
AgaGGCCTACCaTGGtGgTAACGGGTgACGGAGgATTAGGGTTCGATTCCGGAGAGGGAGCCTGAGAAA 70
* 360 * 380 * 400 *
C.vulgaris : CGGCTACCACATCCAAGGAAGGCAGCAGGCGCGCAAATTACCCAATCCTGACACAGGGAGGTAGTGACA :418
D.salina : CGGCTACCACATCCAAGGAAGGCAGCAGGCGCGCAAATTACCCAATCCCAACACGGGGAGGTAGTGACA :419
Fusant : CGGCTACCACATCTAAGGAAGGCAGCAGGCGCGTAAATTACCCAATCCTAACACAGGGAGGTAGTGACA :400
CGGCTACCACATCcAAGGAAGGCAGCAGGCGCGcAAATTACCCAATCCtaACACaGGGAGGTAGTGACA 70
420 * 440 * 460 * 480
C.vulgaris : ATAAATAACAATTACTGGGCCTTTTCAGGTCTGGTAATTGGAATGAGTACAATCTAAACCCCTTAACGA :488
D.salina : ATAAATAACAAT-ACCGGGCATTTTT--GTCTGGTAATTGGAATGAGTACAATCTAAATCCCTTAACGA :487
Fusant : ATAAATAACGAT-GCGGAGCCCTT-CGGGTTCTGCAATCGGAATGAGTACAATTTAAACCCCTTAACGA :468
ATAAATAACaAT aC GgGCctTTtc gGTctgGtAATtGGAATGAGTACAATcTAAAcCCCTTAAC 65
* 500 * 520
C.vulgaris : GGATCAATTGGAGGGCAAGTCTGGTGCCAGCAG :521
D.salina : GTCTCCTTTGGAAGGCAAGTCTGGTGCCAGCAG :520
Fusant : GGATCCATTGGAGGGCAAGTCTGGTGCCAGCAG :501
GgaTCcaTTGGAgGGCAAGTCTGGTGCCAGCAG
Figure 4 Homology region among fusant, D. salina and C. vulgaris
66 Hermin Pancasakti Kusumaningrum & Muhammad Zainuri / Jurnal Teknologi (Sciences & Engineering) 78:4–2 (2016) 61–68
Figure 5 Phylogenetic evolutionary tree displaying the evolutionary relationship of fusant within a lineage shared by the C. vulgaris
species with Dunaliella sp. BBPBAP as an outgroup (str = strain)
Selection on related similarity of fusant with other D.
salina and C. vulgaris species with the
genus Dunaliella, exhibited the 100 and 102 most
related species. Alignment result from selected species
showed that all the species had quite high
percentage of similarity throughout the sequences (81
% to 85 %), presenting such differences inside the
sequences. Analysis by multiple alignment methods
revealed a close relationship of fusant with member of
D. salina and C. vulgaris as illustrated from consensus
tree using phylogenetic evolutionary analisis in Figure 5
and Figure 6. The reliability of the tree topology was
estimated by bootstrapping. The 80 % bootstrap
proportion consensus Neighbor Joining tree for
18SrDNA sequences is shown in Figure 5 and Figure 6.
Homology analysis with C. vulgaris species retrieved
from GenBank is illustrated in Figure 5. which shows
close similarities between a green algae isolates with
those of C. vulgaris strain KMMCC FC-41 and C.
vulgaris strain nm27. According to the phylogenetic
tree, the studied fusant appeared as individual entity
separated from the rest cluster. It was clearly shown
Duna liella sp. BBPBAP Fusant
C. vulgaris str nm27
C. vulgaris CCAP 211/79 C. vulgaris AB080308
C. vulgaris KMMCCFC-1
C. vulgaris strKMMCCFC-15
C. vulgaris str KMMCCFC-42
C. vulgaris strKMMCCFC-12
C. vulgaris str KTP2 C.vulgaris isolate C. vulgaris str CCAP 211/63 C. vulgaris strA1-65
C. vulgaris strCCAP
C. vulgaris strCCAP 211/80
C. vulgaris strLF5
C. vulgaris ccCCAP 211/11P C. vulgaris str
C. vulgaris str: AG-35_ZF1
C. vulgaris strCCAP 211/109
C. vulgaris str IAMC-27
C. vulgaris str: NIES-2172 C. vulgaris var. vulgaris str: NIES-641
C. vulgaris var. vulgaris str: NIES-227 C. vulgaris str: NIES-2170
C. vulgaris var. vulgaris str :NIES-642
C. vulgaris str: NIES-1269
C. vulgaris str: PS-2670
C. vulgaris var. vulgaris strNIES:-
C. vulgaris strKMMCCEC-16
C. vulgaris strKMMCCEC-3
C. vulgaris strKMMCCEC-10
C. vulgaris strKMMCCFC-333
C. vulgaris strSAG 211-11b
C. vulgaris strCCAP 211/110
C. vulgaris strCCAP 211/75
C. vulgaris str
C. vulgaris strCCAP211/8 1 C. vulgaris strCCAP 211/8 2
C. vulgaris strY2
3
671
C vulgaris . strOW01 C .vulgaris strUMTM1
C.vulgaris strKSMA2
C . vulgaris KF574391 C. vulgaris ccKMMCCC-27 C. vulgaris strKMMCCC-111
C. vulgaris ccKMMCCC-88
C. vulgaris strKMMCCC-117
959
669
417
513
C. vulgaris ccKMMCC:FC-
C. vulgaris strKMMCC FC-41
566
1000
0.01
67 Hermin Pancasakti Kusumaningrum & Muhammad Zainuri / Jurnal Teknologi (Sciences & Engineering) 78:4–2 (2016) 61–68
that fusant, instead of having almost equal similarities
with all member of other C. vulgaris, it also had close
relationship with Dunaliella sp. BBPAP as the other
parental.
Figure 6 Phylogenetic evolutionary tree displaying the evolutionary relationship of fusant within a lineage shared by the D. Salina
species. C. vulgaris is used as an outgroup sequence
Homology analysis with D. salina species in Figure
6. showed closest similarities between a green algae
isolates with those of D. salina KU 13. However,
phylogenetic evolutionary tree showed the position
of fusant outside of the cluster of parental. These
results suggested that fusant gained different
character with parental species. In this sense, C.
vulgaris seemed to be more related to D. salina,
which was consistent with homology analysis result.
Almost all of the fusant obtained from protoplast
fusion process suggested higher total carotenoid
production after the process compared with the
parental strain (data not shown).
Although the two profiles of fusant and parental
shared similarity, but the position of fusant in different
cluster with parental indicating that there was indeed
a difference in the cell of the two algal strains. On
the basis of the gained results we could conclude
that the dominant genotype in fusants between D.
salina and C. vulgaris appertain to C. vulgaris. The
difference in the bases profiles is further evidence
that there is indeed a change at the genomic level.
The result also showed that 18SrDNA gene can be
used to calculate dominant genotype in fusant
resulted from protopast fusion process.
4.0 CONCLUSION
Molecular analysis showed that C. vulgaris was more
dominant in fusant comparing with D. salina based
on homology analysis of 18SrDNA sequences. The
result also indicated that the fusant gained both
character from their parents due to conserved
sequence of 18SrDNA in parental and progenitor. The
research showed possibilities in potential acquisition
of genomic combination of both parents.
Acknowledgement
This research was funded by Directorate Research
and Public Services (Ditlitabmas), General of Higher
Education (Ditjen Dikti), Ministry of Research,
Technology and Higher Education, Indonesia year
2015 according to number 023.04.02.1.67345/2015–
Chlorella vulgaris CCAP21111F
D. salina strain
D. salina strain
D. salina strain B34
D. salina str MBTD-CMFRI-S135
D. salina isolate B32
D. salina strain I2
D. salina isolate I3
D. salina strain B33
D. salina strain B25
D. salina strain N5
D. salina strain
D. salina isolate NIOT-28(ANCOST-17)
D. salina strain B24
917
D. salina strain KMMCC
600
D. salina strain KMMCC 1511
428
D. salina strain KMMCC 1064
D. salina str MBTD-CMFRI-S089
D. salina cc DCCBC:CCAP19/20
D. salina strain CS265
D. salina
D. salina strain UTEX LB 200
D. salina strain CCAP
D. salina strain Dsge
D. salina str BuriRam KU01
D. salina strain SAG 19-3
D. salina strain CCAP
D. salina
D. salina strain JR101
D. salina strain JR102
D. salina AF506698
D. salina strain UTEX LB 1644
D. salina strain CCAP
Dunaliella sp. BBPBAP
742
Dunaliella sp. BBPBAP
898
0.01
68 Hermin Pancasakti Kusumaningrum & Muhammad Zainuri / Jurnal Teknologi (Sciences & Engineering) 78:4–2 (2016) 61–68
DIPA 03 Maret 2015 which was gratefully
acknowledged.
References [1] Lee, Y. K. and H. M. Tan. 1988. Interphylum Protoplast
Fusion and Genetic Recombination of the Algae
Porphyridium cruentum and Dunaliella spp. Journal of
General Microbiology. 134: 635–641.
[2] Lu, Y., R. Kong, and L. Hu. 2012. Preparation Protoplasts
from Chlorella protothecoides. World Journal of
Microbiology Biotechnology. 28(4): 1827–1830.
[3] Santiago, C.M. 1982. Protoplast Fusion A New Tecnique for
Genetic Manipulation and Breeding of Industrial
Microorganisme. I.C. Biotech. 5: 435–440
[4] Pulz, O. and W. Gross. 2004. Valuable Products from
Biotechnology of Microalgae. Applied Microbiology
Biotechnology. 65: 635–648.
[5] Borowitzka, M. A. and L. J. Borowitzka. 1988. Dunaliella.
In Borowitzka, M. A. (ed.). Micro-Algal Biotechnology.
Cambridge: Cambridge University Press
[6] Kusumaningrum, H. P. and M. Zainuri. 2014. Optimization
and Stability of Total Pigments Production of Fusan from
Protoplast Fusion of Microalgae Dunaliella and Chlorella in
vivo. Attempts on Production of Sustainable Aquaculture
Natural Food. International Journal of Marine and Aquatic
Resource Conservation and Co-existance. 1(1): 1–5
[7] Guedes, A.C. and F. X. Malcata. 2012: Nutritional Value
and Uses of Microalgae in Aquaculture. In Muchlisin, Z. A.
(ed). Aquaculture. Rijeka: InTech.
[8] Guedes, A. C., H. M. Amaro, and F. X. Malcata, 2011.
Microalgae as Sources of Carotenoids. Marine Drugs. 9:
625–644.
[9] Panaiotov S., Y. Evstatieva, S. Ilieva, V. Levterova, N.
Brankova, D. Nikolova, et al. 2009. Quantitative Assesment
of The Dominant Genome in Fusant Cultures.
Biotechnology and Biotechnological Equipment, supp.1. SE. 23: 892895.
[10] Zainuri, M., E. Kusdiyantini, Widjanarko, J. Soedarsono, and
T. Yuwono, 2003. Preliminary Study on the Use of Yeast
Phaffia rhodozyma as Pigment Source on The Growth of
Tiger Shrimp (Penaeus monodon Fabricius). Indonesian
Journal of Marine Sciences. 8(1): 47–52
[11] Boonyaratpalin, M., S. Thongrod, K. Supamattaya, G.
Britton, and L. E. Schlipalius. 2001. -carotene
Source, Dunaliella salina, and Astaxanthin on
Pigmentation, Growth, Survival and Health of Penaeus
monodon. Aquaculture Research. 32: 182–190.
[12] Kusumaningrum, H. P. and M. Zainuri. Detection of
Bacteria and Fungi Associated with Penaeus monodon
Postlarvae Mortality. International Journal Procedia
Environmental Sciences. 23: 329–337.