Functional Genomics

Yield enhancement is the ultimate goal of crop improvement through genome characterization and manipulation which can be achieved by purposeful genome editing of plant species through alteration of the genetic make by direct gene transformation. Efforts have been made to control insect pests through genetic modification of local cotton varieties through introduction of codon optimized Cry1Ac and Cry2a genes under the control of CAMV35s promoter. Almost 100% mortality of Lepidopteron insects was achieved after 3rd day of detached cotton leaf bioassay when evaluated in lab and showed significant control of these insects in field conditions as well. Similarly, codon optimized cp4 EPSPS transformed in insect-resistant local cotton varieties has shown significant level of glyphosate tolerance after spray at the rate of 1900ml per acre as compared to non-transgenic cotton plants and all grasses which start decaying on 3rd day and showed complete death after 5th day of glyphosate spray assay. Similarly, introduction of different transcription factors, namely WLIM5 in combination with HOX3 and fibre related genes like SUS, AcsA+B, Actin, Aco3 along with pigment structural genes like F3'5'H and DFR flavonoids in different combinations in both Gossypium hirsutum and Gossypium arboreum have resulted in increase of fibre length from 26 mm to 31.5 mm and 18 mm to 26.5 mm respectively also micronaire value was improved from 4.0 to 3.2 and 7.9 to 6.3. Application of molecular breeding and advanced technology like CRISPR CAS 9 system to further enhance the capabilities of the product will be helpful in utilization of full potential of the product developed.

Cotton (Gossypium hirsutum L.) is an important commercial crop that is grown worldwide as a source of fiber and edible oil. Cotton shows higher drought and salt tolerance than do many other major crops such as rice, wheat, and maize. Even so, abiotic stress still have a significant effect on the growth and yield of cotton, which makes it difficult in improvement of salt tolerance in upland cotton. Therefore, the mining of some stress resistance genes will provide potential candidate genes for transgenic resistance breeding in cotton. Some salt stress-related gene high-affinity potassium transporter play an important role in plant salt resistance pathway. Meanwhile, a putative high-affinity potassium transporter gene SbHKT was isolated from halophyte Salicornia bigelovii by RACE cloning in an earlier research. We conducted a preliminary functional analysis by transferring the gene into tobacco and cotton with agrobacterium-mediated transgenic technology. By the use of Agrobacterium tumefaciens high-affinity K+ SbHKT gene was transferred into tobacco, and resistance plants were screened. By PCR detection of T0 generation regenerated plants, transgenic tobacco plants were successfully expressed at the mRNA level. Identification salt tolerance of transgenic tobacco indicated the transgenic plants were more resistant to the salt tolerant. SbHKT gene was transportered into cotton, and southern blot analysis of positive plants shows the gene SbHKT has been integrated into the cotton genome. Tissue-specifc expression showed that SbHKT is expressed at differential levels in all tissues examined and strongly induced by various phytohormones and abiotic stress. In vivo and in vitro subcellular localization suggested that SbHKT is located in the plasmamembrane. In response to salt stress, transgenic cotton plants overexpressing SbHKT showed signifcantly higher germination rates, longer roots, and more vigorous growth than wild-type plants. These findings demonstrated that SbHKT plays an important role in the abiotic stress response, and that overexpression of SbHKT in transgenic cotton improves salt tolerance.

Cotton fibres arise from the epidermal cells of the seed coat and may be either long (lint) or very short (fuzz). Both lint and fuzz are single-celled and indistinguishable in appearance during the early stages of their growth, suggesting that their growth may involve the same physiological and biochemical processes that are regulated by the same set of genes. The dominant fuzzless mutation N1 of tetraploid Gossypium hirsutum has been demonstrated to be a defective allele of the At-subgenome homoeolog of MYB25-like, a master gene regulating fibre initiation as silencing MYB25-like resulted in lintless and fuzzless seeds. We recently identified five genetic loci, including a major contributing locus containing MYB25-like_Dt, associated with the recessive fuzzless seed trait n2 in G. barbadense based on genotyping (SNP chip and mapping-by-sequencing) of fuzzy and fuzzless near isogenic lines (NILs) derived from an interspecies cross (G. barbadense x G. hirsutum). We compared the expression changes of MYB25-like_At and MYB25-like_Dt during fibre development in cotton accessions with different fibre phenotypes. At 3 dpa (days-post-anthesis) when fuzz fibres are initiating, expression of MYB25-like_Dt was significantly lower in fuzzless NILs than in fuzzy seeded NILs, while higher MYB25-like_Dt expression was associated with more seed fuzz across different cotton genotypes. Phenotypic and genotypic analysis of MYB25-like homoeoalleles in cottons showing different fibre phenotypes and their crossing progeny indicated that both MYB25-like_At and MYB25-like_Dt are associated with lint development, and that fuzz development is mainly determined by the expression level of MYB25-like_Dt at ~3 dpa, suggesting that cotton lint and fuzz development are regulated by the temporal expression of MYB25-like homoeologs. In this presentation, we will propose a working model for the role of MYB25-like homoeologs in lint and fuzz development, and discuss strategies for confirmation of the contributions of MYB25-like homoeologs to lint and fuzz development.