Working group session:
Functional Genomics
Presentation type:
poster
Authors:
Qayyum Rao, Abdul; Husnain, Tayyab
Presenter:
Qayyum Rao, Abdul
Correspondent:
Qayyum Rao, Abdul
Abstract:
Begomoviruses are known to be a major problem in the cotton and other crops in Pakistan particularly in the Punjab region. Crops infected with these viruses show a notable decrease in the yield which results in major losses for farmers in particular and for the nation’s economy in general. Genetically-engineered resistance against begomoviruses is at advanced stage of investigation and crops such as cotton are more likely to benefit of solving this problem.
In current study a collaborative effort has been made by different institutes namely IAGS, NIBGE and CEMB with financial support of USDA. In this effort strategies employed includes nine construct namely C1 to C6 of which C1 was based on pathogen derived resistance against vector, three constructs from C2 to C4 were based upon pathogen derived resistance against Gemini viruses while remaining two constructs i.e C5 and C6 were developed on the principle of non-pathogen derived resistance by NIBGE i.e protein mediated resistance against CLCuV were handed over to CEMB for transformation Similarly One construct of pathogen derived resistance against Gemini viruses (RNAi based construct against sense and anti-sense region of replication of begomoviruses) namely C7, other RNAi based constructs against beta satellite of CLCuV namely C8 and one RNAi based construct against cathepsin gut region of whitefly namely C9 was developed by IAGS and provided to CEMB for transformation in local cotton varieties.
A local modified gene transformation technology in local cotton cultivar namely MNH-786, VH-289 and CIM-496 has been applied with increased efficiency. Multiplication of putative transgenic cotton plants has been done in controlled condition of green house after one and half month screening on selection medium. Generation advancement and screening of best event from T0 to T1 and T2 of transgenic cotton plants was done and also in process. Confirmation of successful integration of transgene was done through amplification by using gene specific primers for C1, C2, C3, C4, C5, C6 and C7. Quantification of miRNA in transgenic cotton plants was done on relative basis through real time PCR in both To and T1 generation. Variable quantity of miRNA was found in different events in both To and T1 generation of transgenic cotton plants. Morphological characteristics of these transgenic plants in comparison of miRNA quantity have shown that CLCuV symptoms were less in plants having greater quantity of miRNA and greater in plants having less quantity of miRNA. Similarly absolute quantification of virus titer was done in nine C2 transgenic plants. Four transgenic plants showed least quantity of virus with no morphological symptoms as compared to other five which have shown high titer of virus with maximum viral symptoms.
Further generation advancement and selfing of these transgenic cotton plants with determination of their copy no and virus titer will be helpful for the selection of best events which will be expected outcome of this study in near future.