To express selected coconut genes associated with oil biosynthesis in corn (Zea mays L.)
All activities concerning gene cloning, construction of transformation cassettes and tissue culture will be conducted inside a BSL2 facility. Likewise Seedling will be regenerated inside a BL2 greenhouse. Access to both facilities will be restricted to authorized personnel only. All materials that will be used during and after each experiments will be sterilized and decontaminated using necessary procedures. All personnel that will be involved in the project will be informed and will adhere to Good Laboratory Practices (GLP)
a) Activities shall be conducted at the P2 - Biochemistry Laboratory of the University of the Philippines Los Baños - Institute of Plant Breeding.
b) The proponent shall adhere as closely as possible to the schedule of activities reflected in the submitted Gantt chart.
c) Any modifications in the schedule of activities shall be made with the concurrence of the UPLB-IBC and the DOST-BC.
d) Signage should be in place indicating that the facility is restricted/limited access.
e) The proponent must provide a logbook to record access of personnel to the experimental facility.
f) The proponent must take measures to ensure that only the DOST-BC authorized personnel are allowed inside the experimental facilities.
g) The experimental facilities must be kept under lock when not in use.
h) In case of undue destruction of experimental materials resulting from unauthorized entry of personnel or breach of containment of experimental facilities, the proponent shall implement measures to prevent the inadvertent escape of any viable material within the facility.
i) The proponent and the supervising IBC shall be held accountable for the undue destruction of the experimental materials and the consequences that their inadvertent escape may cause to the surrounding environment.
j) The DOST-BC should be informed in advance of any visitations by unauthorized persons.
k) The DOST-BC shall be informed immediately of any intrusion by unauthorized persons.
l) The proponent shall strictly observe proper disposal procedures for all materials.
m) Movement of all materials will be done in compliance with all relevant biosafety and phytosanitary requirements of the Philippines.
n) Any additional requirement that the DOST-BC may impose, as necessary, for the duration of the experiment shall be complied with.
o) The proponent shall submit through the IBC a completion report within 90 days after completion of the experiment.
p) The IBC shall submit to the DOST-BC a report on the completion of this project, in the prescribed format.
One of the major goals of the agricultural sector worldwide is to meet the unceasing rise in demand for oil consumption. Studies have shown that Wrinkled1 gene (WRl1) act as master regulator for plant oil biosynthesis and alteration in its expression may result in the improvement of oil content in crops. In this project, we would like to express WRI1 in an experimental monocot model system, corn. The overarching goal of this activity is to demonstrate that oil biosynthesis related genes could be effectively expressed through the use of an in-house transformation system and in turn provide a proof of concept strategy prior to moving onto monocot crops with longer developmental cycles.
In order to meet the aforementioned goal, a three and a half year project that is currently being implemented at the Institute of Plant Breeding, UPLB is accomplishing the following: isolate, clone and characterize wrinkled 1 from solid endosperm of coconut, construct transformation cassette with coconut wrinkled 1 (CnWRl1), determine yellow corn inbered lines that will be suitable for the transformation activity, transform cassette containing CnWRl1 in the chosen yellow corn inbred lines using particle inflow gun and regenerate transformed corn tissues into whole plants under contained laboratory conditions.
Currently, an ortholog of Zea mays Wrinkled (WRl1) gene derived from coconut (CnWRl1) has been engineered into a plant expression vector, pCAMBIA1302_CnWRl1. Another 2 expression vectors containing reporter genes have been secured from the Plant Laboratory of the NIMBB, Diliman (i.e Pcambia1302 with mGFP5 and PBl121_DsRed with DsRed). Likewise, two yellow corn inbreds, CML161 and Pl123 have been determined as the most suitable lines transformation.
Using a fabricated particle inflow gun, the contructed vector containing CnWRl1 will be cotransformed with either one of the vectors containing a reporter gene into immature zygotic embryos and embryogenic calli. Success of transformation will be determined using flourescence microscopy and antibiotic selection. Expression analysis will be done once regenerants are formed using qRT-PCR.
Public Information Sheet
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