Simposio sobre Evaluación de Riesgo Ambiental
México, D.F., Marzo 2-3, 2011
Estudio de caso No. 1: Frijol común
resistente a virosis
Francisco J. L. Aragão
Embrapa/Cenargen
[email protected]
Brazil 3.4 M ton – 4.1 M ha
Mexico 1.1 M ton – 1.4 M ha
Diseases (golden mosaic, white mold)
Drought
Insects (field and post-harvest losses)
Economics
Annual reductions in the range of 90,000 to 280,000 tons
Enough to feed 6 M to 20 M adults.
180,000 ha are unsuitable
yellow-green mosaic of leaves, stunted growth
distorted pods and seeds
RC
RC
AC1
DNA-A
AV1
DNA-B
BC1
AC2
AC3
BGMV is transmitted by the whitefly Bemisia
tabaci in a persistent, circulative manner
BV1
Bean golden mosaic
Identity PvAhas=83%
Catalytic sites (ThDP)=98-100%
2/22
Embrapa 5.1
EMB-PVØ51-1
MPMI, 2007
Nature Biotechnology, 2009
CTNBio NORMATIVE RESOLUTION NO. 05
MARCH, 2008
Gives provisions on rules for commercial release of
Genetically Modified Organisms and their derivatives
Portuguese:
http://www.ctnbio.gov.br/index.php/content/view/114
44.html
English:
http://www.ctnbio.gov.br/index.php/content/view/128
57.html
Common bean
Embrapa 5.1
EMB-PVØ51-1
Embrapa Genetic
Resources
and Biotechnology
Embrapa Rice and Bean
Embrapa Maize and
Shorgum
Embrapa Agrobiology
Embrapa Agroindustry
UNESP-Botucatu
Embrapa Soybean
Antonio Félix da Costa, F. Aragão
Molecular characterization
•
•
•
•
•
•
•
Number of DNA inserts, insert stability
Number of copies of genetic elements
within the insert
Integrity of gene cassettes
Presence of additional DNA (backbone)
Sequence of genomic flanking DNA
Sequence of the inserted DNA
DNA LandMarks
...
...
No functional bla gene
Single locus: segregation of: 3:1
Stability
Generations x after sexual crosses
Agronomic equivalence,
Environmental safety
•Agronomic / phenotypic assessments
•Weediness assessment (HT)
•Fitness
•Environmental safety
•Environmental fate
mobio+Fast Spin
Fast Spin
100
90
80
70
60
50
mobio
40
30
20
10
0
casa de vegetação
Feijão OLATHE 5.1
15 dias
2
Feijão OLATHE 5.1
15 dias
3
Feijão OLATHE
15 dias
7
Feijão OLATHE 5.1
15 dias
1
Feijão OLATHE
15 dias
8
Feijão OLATHE
15 dias
5
Feijão OLATHE
15 dias
6
Feijão OLATHE 5.1
15 dias
4
Feijão OLATHE 5.1
15 dias
5
Feijão OLATHE 5.1
15 dias
6
Feijão OLATHE
15 dias
2
Feijão OLATHE
15 dias
3
Feijão OLATHE
15 dias
4
Feijão OLATHE
15 dias
1
No consistent change of bacterial community based on 16s rDNA
Pests
Predators
Densidade de esporos de FMAs
(nº por 50 mL de solo)
35
Goiânia - 2008
28
21
14
7
0
Olathe Pinto
Olathe 5.1
Variedades de Feijão
Arthropods
Mycorrhizal fungi
160
Olathe Pinto convencional
Olathe Pinto transgênico
150
140
2008
120
110
100
90
80
40
40
40
70
30
20
10
5/0
8
20
10
0
28
/0
21
/0
5/0
8
5/0
8
14
/0
5/0
8
07
/0
30
/0
4/0
8
4/0
8
23
/0
16
/0
4/0
8
0
30
a
a
Olathe Pinto
Olathe 5.1
Densidade de esporos de FMAs
(nº por 50 mL de solo)
40
Densidade de esporos de FMAs
(nº por 50 mL de solo)
50
Sete Lagoas - 2008
Goiânia - 2008
Londrina - 2008
60
Densidade de esporos de FMAs
(nº por 50 mL de solo)
Número médio de indivíduos
130
30
20
10
0
Varie dade de Fe ijão
a
a
Olathe Pinto
Olathe 5.1
30
20
10
0
Varie dade de Fe ijão
a
a
Olathe Pinto
Olathe 5.1
Varie dade de Fe ijão
Data
110
100
90
80
70
60
50
300
200
100
0
40
Densidade de esporos de FMAs
(nº por 50 mL de solo)
120
a
a
Olathe Pinto
Olathe 5.1
Varie dade de Fe ijão
30
300
200
100
0
a
a
Olathe Pinto
Olathe 5.1
300
200
100
Varie dade de Fe ijão
20
10
Data
/09
20
/05
/09
13
/05
/09
06
/05
/09
29
/04
/09
22
/04
/09
15
/04
/09
0
08
/04
Número médio de indivíduos
130
Sete Lagoas - 2009
Goiânia - 2009
Londrina - 2009
Densidade de esporos de FMAs
(nº por 50 mL de solo)
140
2009
400
400
400
Olathe Pinto convencional
Olathe Pinto transgênico
150
Densidade de esporos de FMAs
(nº por 50 mL de solo)
160
Biochemical analysis
0
a
a
Olathe Pinto
Olathe 5.1
Varie dade de Fe ijão
50
70
65
60
48
55
50
Olathe 5.1 transgênico
Olathe convencional
Jalo Precoce
Pérola
Arcelina
46
45
40
44
35
30
36
25
Olathe 5.1 transgênico
Olathe Convencional
Jalo Precoce
Pérola
Arcelina
15
10
5
Fêmeas
0
Número médio de adultos
65
60
55
50
45
40
35
30
25
Olathe 5.1 transgênico
Olathe Covencional
Jalo Precoce
Pérola
Arcelina
20
15
10
5
Machos
0
34
32
Período médio de desenvolvimento (dias)
20
30
Fêmeas
B
1
2
3
4
5
6
7
48
46
44
10
34
32
30
Machos
C
1
2
3
4
5
6
7
4
5
6
7
48
100
46
90
9
36
120
110
8
Olathe 5.1 transgênico
Olathe convencional
Jalo Precoce
Pérola
Arcelina
80
8
9
10
Olathe 5.1 transgênico
Olathe convencional
Jalo Precoce
Pérola
Arcelina
44
70
60
36
50
40
Olathe 5.1 transgênico
Olathe Convencional
Jalo Precoce
Pérola
Arcelina
30
20
10
Fêmeas + Machos
34
32
30
Fêmeas + Machos
0
1
2
3
4
5
6
Geração
7
8
9
10
1
2
3
8
Geração
Effect on Zabrotes subfasciatus for several generations
9
10
Nature Biotechnology, 2009
Composition equivalence,
Nutritional equivalence
•Grain samples from multi-location, multi-year,
replicated field trials
•Secondary metabolites
•Pilot scale processing, nutrient / antinutrient analysis
(validated methods)
•Confirmation of food/feed safety
•Animal feeding studies
Sugars: Sucrose, raffinose, Stachyose
Vitamins: Vitamin B1, Vitamin B2
Amino acids: Tryptophan, Cysteine, Methionine,
Glutamic Acid, Serina, Glycine, Histidine, Arginine,
Threonine, Alanine, Proline, Tyrosine, Valine, Lysine,
Isoleucine, Leucine, Phenylalanine.
Physic-Chemical Analyses: Moisture, ash, protein,
phytic acid, trypsin inhibitor, etc
Minerals: Aluminum, Calcium, Lead, Cobalt, Copper,
Chromium, Iron, Phosphorus, Magnesium, Molybdenum,
Potassium, Selenium, Sodium, Zinc
Sensorial Analyses
1: Arcelin-5B. Chain: 22-261, pI: 6.24, Mw: 27055
pH 4
7
kDa
161718
66
13 15
14
12
2: Glutelin type-A 1 acidic chain. Chain: 25-306, pI: 6.59, Mw: 3189
3: Isoform 2 of 12S seed storage protein CRA1 OS=Arabidopsis... Chain: 1-368, pI:
6.62, Mw: 41032
21 20 19
4: Arachin 21 kDa protein. Chain: 1-176, pI: 4.29, Mw: 20863
25 26
5: Arachin 21 kDa protein. Chain: 1-176, pI: 4.29, Mw: 20863
2423
22
45
6: Arcelin-5B. Chain: 22-261, pI: 6.24, Mw: 27055
3
7: Arcelin-5B. Chain: 22-261, pI: 6.24, Mw: 27055
30
2 1
9
8: Arachin 25 kDa protein. Chain: 1-201, pI: 7.12, Mw: 22220
7
6
9: Glycinin A2 subunit. Chain: 19-296, pI: 4.90, Mw: 31623
10: Conglutin. Chain: 22-145, pI: 5.50, Mw: 14846
8
20
11: Conglutin. Chain: 22-145, pI: 5.50, Mw: 14846
12: Beta-conglycinin, alpha chain. Chain: 63-605, pI: 4.87, Mw: 63152
10
13: Beta-conglycinin, alpha chain. Chain: 63-605, pI: 4.87, Mw: 63152
11
14: Beta-conglycinin, alpha chain. Chain: 63-605, pI: 4.87, Mw: 63152
5 4
14.4
15: Beta-conglycinin, alpha chain. Chain: 63-605, pI: 4.87, Mw: 63152
16: Glutenin, high molecular weight subunit PW212. Chain: 22-838, pI: 5.38, Mw: 87008
17: Glutenin, high molecular weight subunit PW212. Chain: 22-838, pI: 5.38, Mw: 87008
18: Glutenin, high molecular weight subunit PW212. Chain: 22-838, pI: 5.38, Mw: 87008
19: Convicilin. Chain: 29-571, pI: 6.11, Mw: 63932
20: Convicilin. Chain: 29-571, pI: 6.11, Mw: 63932
21: Convicilin. Chain: 29-571, pI: 6.11, Mw: 63932
22: Phaseolin. Chain: 25-428, pI: 5.25, Mw: 45383
23: Phaseolin. Chain: 25-428, pI: 5.25, Mw: 45383
24: Phaseolin. Chain: 25-428, pI: 5.25, Mw: 45383
25: Phaseolin, alpha-type. Chain: 25-436, pI: 5.13, Mw: 46698
26: Phaseolin, alpha-type. Chain: 25-436, pI: 5.13, Mw: 46698
Rattus norvegicus
siRNA and grains
180
P-i
P-f
160
140
120
100
80
60
40
20
0
GC
GFT
GFC
GA
Biochemical Analysis
urea, creatinine for assessment of
renal dysfunction; ALT-SGPT (alanine
aminotransferase - glutamicopivic
transaminase) and AST-SGOT
(aspartate aminotransferase aspartate aminotransferase) to
assess liver injury; alkaline
phosphatase (ALP) and total serum
protein (PTs ) to evaluate liver
dysfunction
Stomach, intestine,
duodenum, jejunum, uterus,
hepatocytes, liver, kidney
Post-commercial monitoring – 5 years
Embrapa Recursos Genéticos e Biotecnologia
Francisco José Lima Aragão
Elsa Oliveira Paranaguá e Lago Nogueira
Kenny Bonfim
Maria Laine Penha Tinoco
Antonieta Nassif Salomão
Solange Carvalho Barrios Roveri José
Marcelo Porto Bemquerer
Beatriz Simas Magalhães
Vera Lucia Perussi Polez
Embrapa Arroz e Feijão
Josias Corrêa de Faria
Eliane Dias Quintela
José Francisco Arruda e Silva
Edmar Cardoso de Moura
Vanderlino Moreira de Santana
Jaison Pereira de Oliveira
Murillo Lobo Junior
Paula Arielle Mendes Ribeiro Valdisser
Maria José Del Peloso
Embrapa Agroindústria de Alimentos
José Luiz Viana de Carvalho
Marília Regini Nutti
Edson Watanabe
Edna Maria Morais Oliveira
Ronoel Luiz de Oliveira Godoy
Sidinea Cordeiro de Freitas
Sidney Pacheco
Luzimar da Silva de Mattos
Jeane Santos Rosa de Mello
Manuela Cristina Pessanha de Araujo
Adriana Paula da Silva Minguita
Carmine Conte
Epaminondas Silva Simas
Juliana de Oliveira Santos
Tania dos Santos Silva
Jose Manoel de Oliveira
Paulo Sergio de Souza
Tatiane Correa de Oliveira
Embrapa Agrobiologia
Bruno José Rodrigues Alves
Gustavo Ribeiro Xavier
Segundo Sacramento Urquiaga Caballero
Altiberto Moreira Baeta;
Roberto Gregio de Souza;
Maria Elisabeth Fernandes Correia
Norma Gouvea Rumjane
Roberto Silva de Oliveira
Itamar Garcia Ignácio
Orivaldo José Saggin Junior
João Luiz Bastos
Universidade de Brasília
Élida Geralda Campos
Anna Paula Costa Jesuino
Érica Heringer Machado
Viviane Yllena Vieira de Souza
Universidade Estadual de Campinas
Jaime Amaya-Farfan
Embrapa Milho e Sorgo
José Aloisio Alves Moreira
Embrapa Soja
Geraldo Estevam de Souza Carneiro
Universidade Estadual Paulista
Norka Beatriz Barrueto González
Renata M. Galvão Campos Cintra
Luis Fernando Barbisan
Alaor Aparecido de Almeida
Universidade Federal do Ceará
Francisco de Assis de Paiva Campos
Muchas
Gracias !