BIOSAFETY SERTIFICATION GENETICALLY MODIFIED PLANTS...
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BIOSAFETY SERTIFICATION GENETICALLY MODIFIED PLANTS : Drought Tolerance GM Sugarcane A Case Study Nurmalasari Darsono Biology Dept, FST, Airlangga Unversity International Seminar and Workshop on Biotechnology and Biosafety Jember University, 10-12 July, 2019
Transcript of BIOSAFETY SERTIFICATION GENETICALLY MODIFIED PLANTS...
BIOSAFETY SERTIFICATIONGENETICALLY MODIFIED PLANTS :Drought Tolerance GM Sugarcane
A Case Study
Nurmalasari DarsonoBiology Dept, FST, Airlangga Unversity
International Seminar and Workshop onBiotechnology and Biosafety
Jember University, 10-12 July, 2019
THE BENEFITS OF SUGARCANE
Paper based products
BagasseMolasses
Sugar wax
Sugar (crystals of sucrose)
Bio-ethanol
Sugarcane
Sugarcane productivity has decreased due to increasing usage of dry land.
Dry land characteristics : water deficient, high evaporation rates, high day temperature during summer, low humidity, and high run off and soil erosion, saline soil and low in fertility.
INTRODUCTION
INTRODUCTION
Gambar 2. Trend Peningkatan Areal Lahan Kering / Tegal di PTPN XI(Sumber : Bidang Tanaman PTPN XI)
76.615,7 64.513 59.884,2 65.719,1 68.602,,90
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2008 2009 2010 2011 2012
PR
OS
EN
TA
SE
TAHUN
TREN PENINGKATAN AREAL LAHAN KERING/TEGAL
LAHAN TEGAL
LAHAN SAWAH
Gambar 3. Produktivitas per Kategori Lahan(Sumber : Bidang Tanaman PTPN XI)
INTRODUCTION
CONVENTIONAL BREEDING
BIOTECH BREEDING
Limiting Factors of Conventional Breeding
1. Sugarcane pollination in natural condition is very rare :
• there’s no animal spreading pollen,
• different time of flower maturity
• low viability of pollen
2. Sugarcane seeds are very small, low viability (only few minutes), have no dormancyperiode, and cannot be grown on land naturally
Developing New Varieties
1. Gene Technology has been considered as a powerful method to anticipate a shift in planting sugarcane to dry land, addressing issues of water availability and climate change.
2. Specific character can be introduced into plant tissue through genetic engineering
3. Cooperations with other institutions (foreign & domestic) are needed
4. precise, rapid, efficient and cost effective
PTPN XI Strategy
for Developing New Varieties through Biotechnology
THE SELECTION OF COMMERCIAL SUGAR CANE VARIETIES AS PARENTAL
DROUGHT TOLERANCE, HIGH SUGAR CONTENT SUGARCANE
EXAMINATION OF MOLECULAR BIOLOGY AND PHISIOLOGY CHARACTER
SUGARCANE TISSUE CULTURE
GENETIC TRANSFORMATION
FASILITAS UJI TERBATAS (LFT) DAN CONFINED FIELD TRIAL (CFT)
FOOD SAFETY AND ENVIRONMENT SAFETY ASESSMENT (PP 21 TH. 2005)
Releasing New Variety
GM Sugarcane using betA gene encoding choline dehydrogenase
Choline dehy-drogenase (CDH)
Choline mono-oxygenase (CMO)
Bacteria
Plants
Betaine aldehyde dehydrogenase (BADH)
Compatible solute
Synthesis of choline and GB in plants.
Some plants can synthesize GB.
Synthesis of glycine betaine(GB) from choline in bacteria
a. Parental : BL (Bulu Lawang)
b. Character : betA gen coding for protein choline
dehydrogenase (CDH). betA gen was taken from
Rhizobium meliloti (soil bacteria) by Ajico, Intl. cloned to
pMLH213 with Agrobacterium tumefaciens strain LB 4404
as host. Genetic transformation (biological method) to
sugarcane by PT. Perkebunan Nusantara XI
LB PNOS NPT II TNOS E1E2 P35S betA TNOS P35S HPT T35S RB
XbaI SacI
pMLH2113-betA
Eksplancallus Seleksi Regenerasi
Peta konstruk plasmid rekombinan pMLH2113-betA
Tahapan transformasi genetik metode biologismelalui Agrobacterium tumefaciens
Gambar 10.Plantlet BL dalam media dengan konsentrasi NaCl (dari ki-ka : 0,3; 0,25; 0,20; 0,15; 0,1; 0) pada harike-21 (minggu ke-3)
Gambar 11.Plantlet NXI-4T dalam media dengan Konsentrasi NaCl (dari ki-ka) 0,30; 0,25; 0,20; 0,15; 0,10 ; 0 ) pada hari ke-21 (minggu ke-3)
(Sumber data : Laboratorium Bioteknologi PTPN XI)
Tolerance to osmoticstress
Growth response in water deficiency
Sugarcane Cultivars
Lenght of water stress (days)
Wilting Permanent Wilting
Dried
NXI-3T 12 > 30 > 30
NXI-4T 19 > 36 > 36
NXI-5T 14 > 30 > 30
BL579-NT(Control)
6 19 19
PSJT 941(non GM drought tolerant cane)
19 > 36 > 36
Uji kering dilakukan dgn penghentian pemberian air
Respon 8 hari kekeringan, dari kanan : kontrol, 3T, 4T, 5T
Profil akar –tebu non PRG
Profil akar -tebu PRG NXI-4T
Profil akar –PSJT 941
ADMINISTRATOR:
Biosafety Commission for GM Product (byPresident Regulation) and its organs :
1. Environment Safety Division
2. Food Safety Division
3. Feed Safety Division
4. Assessment team of social economicimpact (2012)
BASIC REGULATION :1. Laws of the RI no. 21/2004 : Ratification of
Cartagena Protocol about biosafety for biodivesity convention
2. Govt regulation no. 21/2005 : Biosafety of GM Product
3. Minister of Agriculture regulation no. 61/ Permentan/OT.140/10/2011 : Examination, assessment, release and discharge of plant varieties (Website Indonesia Biosafety Clearing House : indonesiabch.or.id)
Environment Safety AssesmentGenetic Stability
Genetic expression
Plant performance
Study of gene flow, displacement
Study of invasiveness
Study of non target organism
1. Molecular testing (using Southern Blott)
2. Biochemistry testing (using HPLC)
3. Physiologycal Character test (NaCl test)
4. Morphology character (shoot, root, nodes length)
Pollen and Seed
Soil Bacteria
Soil Parameter
Diversity of Soil Microbe
LIMITED FIELD TRIAL
Definition :A study area of genetically modified
plants that require restrictive measures against planting material
and new gene in order to remain in its study site.
LFT is a mandatory if GM plants intended to be released commercially
Permentan no. 61/2011
Genetic stability and Gene Expression
1T 4T 6T NT
betA
CaMV
PCR analysis for inserted gene detection (3 vegetative generation)
Detection of inserted gene (Southern Blot method)
Data:03061017.D01 Method:03061017.M01 Ch=1Chrom:03061017.C01 Atten:6
0 5 10min
0
20
40
60
mAbs
2.375
5.324
6.068
7.042
7.877
9.662
10.408
10.825
Betain Standard
Data:07061011.D01 Method:07061011.M01 Ch=1Chrom:07061011.C01 Atten:6
0 2 4 6 8 10min
0
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60
mAbs
2.289
4.598
5.322
5.726
9.529
Betain in sugarcane juice
Betaine content on sugarcane juice (HPLC method)
Layout LFT
Main field road as control pathway
LIMITED FIELD TRIAL
Adjacent field with different period of planting
Outside drainage
Uncultivated row
LFT GM sugarcane
Distance isolation to avoid contamination or gene flow
