G. Arinaitwe, H. Khana, P. Namanya, . Kiggundu, A W.K Tushemereirwe and J. Dale Research Framework National Research Laboratories - Kawanda Genetic modification of highland bananas: biofortification with PVA and iron
Nov 29, 2014
G. Arinaitwe, H. Khana, P. Namanya, . Kiggundu, A W.K Tushemereirwe and J. Dale
Research Framework
National Research Laboratories - Kawanda
Genetic modification of highland bananas: biofortification with PVA and iron
Introduction
�Uganda has a population of 34 M people & is projected to be 53 M by 2025
Problems:
�Inadequate food quantities
�Imbalanced nutrients,
containing mainly
carbohydrates
Banana is synonymous to food
A B C
Bananas in Uganda
Banana weevils
Nematode effect
Banana bacterial wilt
Black Sigatoka disease
Micronutrient deficiencies in Uganda
Micronutrient undernutrition
� The major micronutrient deficiencies in Uganda include:�Vitamin A deficiency (VAD) �Iron deficiency Anaemia (IDA) �Iodine Deficiency Disorders (IDD)
� National Nutritional Surveys 1994-1999�31 out of 36 districts had VAD�50% of children have VAD�40% of children > 5 years have IDA�50% of pregnant women had IDA�30% of maternal deaths were attributable to IDA
psyB73
crt I
psy2 Psy2a
psy
Sources of genes for biofortification
Constructs used in biofortification
1st Batch 2nd Batch(Opt series)
3rd Batch(GEN series)
E1-ZmPsy1 E1-APSy2a E1-ZmPsy1Q60
E1-Crt1 Acs-ZmPsy1Q60+E1-S.Fer E1-APsy2a
E1-APsy2a Acs-ZmPsy1Q60 BT4act-APsy2a
E1-S.Fer E1-ZmPsy1Q60 Ubi-APsy2a
35S-Fro2 E1-ZmPsy1 Ubi-ced9
35S-IrtI Ubi-mced9 Ubi-AtBag4
Ubi-Gus
E1-Gus
Biofortified product for low and highlands
Cultivars: ‘Nakinyika’ (EA-AAA)And ‘Sukali Ndizi’ (ABB)
Cultivars: Hybrid M09Tolerant to: Black Sigatoka, Nematodes and weevils
ECSs work at Kawanda, Uganda
8. Weaning and potting of plantlets
2. Inoculation stage
3. Embryogenic callus
7. Multiple shoot formation and rooting
4. Cell in suspension
5. Somatic embryos
formed from suspensions
1. Male bud
6. Embryo germination
Genetic transformation
Materials and methods
ECS initiation and transformation
ECSs
Selection
Embryogenic callus
ECS infection
Shoots from EC clones
PCR analysesMorphological analyses Southern blot analyses
Male bud Immature flower
Future activities
NARO-KARI Biotechnology Laboratory
Shoot regeneration improved
0.5 mg/L GA3 1.0 mg/L GA3 1.5mg/L GA3Control: 0mg GA3
2 Weeks 1.5 Months 2.5 MonthsTra
nsf
orm
atio
nan
d s
elec
tio
n
Shoot regeneration
Weaning and hardening
First CFT at NARO, Uganda
CFTs at NARO-Kawanda
� Several CFTs of transgenic lines containing disease resistance genes and micronutrient traits
Phenotypes observed
Cultivar %Water contentBCE (µg/g DW)
Day1 Day7 Day 14
Mbwazirume 76.3 9.8 17.5 23.5
Nakinyika 73.9 13.5 21.0 26.5
Nakitembe 76.9 19.6 23.4 28.3
Kisansa 75.1 9.1 18.0 18.8
M9 74.5 4.3 11.0 16.8
Butobe 58.2 18.2 22.3 23.1
Kayinja 65.2 0.8 1.7 2.2
Carotenoid content at FG during storage
NAKINYIKA HYBRID M9 KAYINJANAKITEMBE
High carotenoid content in transgenic S. NdiziHigh carotenoid content in transgenic S. Ndizi
1
2
3
4
Control line ZmPsy1 (12-39) lineCross section of the control and the ZmPsy1 (12-39) line; above: Chromatogram peaks for 12-39 fruit extract identified at λmax=450nm, peak 1, lutein at 6min; peak 2, α-carotene at 12.2; peak 3, trans-β-carotene 13.1min & peak4, cis β-carotene at 13.9min
PVA retention in target cooking bananas
Steamed
Boiled
FOC symptoms of the diseaseYellow leaves
Discoloured pseudostem
Discoloured corm
� Microconidia entry to the corm is through wounded /cut root hairs
� The conidia spreads up the xylem and blocks water and nutrient movement
� Death of plant releases the dormant chlamydospores back into the soil
� Reinfection is through running water and propagating suckers
ND control ND 1 ND 2 ND 3 ND 4
ND control ND 1 ND 2 ND 3 ND 4
FOC wilt in cultivar Sukali Ndizi
1212 55 72 83 NTC
Acknowledgements
NARO
A. Kiggundu G. Arinaitwe
P. LamwakaB. Magambo
R. Ecodu
P.Namanya
QUT
Rob HardingDoug Becker
Marion BatesonJason Geijskes
Harjeet KhannaAnthony James
Jen Kleidon
Bulu Malazi
Prof James Dale (Program Leader)
Dr. WilberforceTushemereirwe