The availability of reproducible regeneration system through tissue culture is a major bottleneck in wheat improvement program. The present study has considered to develop an efficient callus induction and regeneration system using mature and immature embryos as explants in recently released agronomically superior spring wheat varieties. An efficient sterilization process was standardized using 0.1% HgCl and 70% ethanol for both seeds and embryos. The maximum possible combinations of plant growth regulators (PGRs) were evaluated for their effect on different wheat regeneration processes through tissue culture starting from callus to root induction. Picloram is found as an effective auxin with 87.63-98.67% callus induction efficiency in both explants. Supplementation of CuSO along with 2,4-D, zeatin in regeneration medium significantly enhanced the multiple shoot induction. The shoot development was achieved using full strength Murashige and Skoog's (MS) medium and root induction using half MS medium without PGRs. The optimized medium and method has resulted up to 100% regeneration irrespective of the genotype used with high reproducibility. Thus, the standardized regeneration system can be used in the regeneration of healthy plants from embryos rescued from interspecies crosses, transgenic production, induced mutation breeding and recently developed genome editing techniques for the procreation of wheat plants having novel traits.

Collections of Puccinia triticina were made from farmers’ fields of five different agro-ecological locations (Sakrand, Tandojam, Larkana, Sanghar and Badin) of Sindh province, Pakistan from 2015 and 2016, to identify the virulence variation. Single uredinial isolates were investigated for virulence phenotyping on 24 near isogenic (Thatcher wheat) lines which differ for single Lr resistance genes. Spores from two locations (Sakrand and Tandojam) were not viable and could not be revived and only urediniospores of three locations (Larkana, Sanghar and Badin) were revived. None of the pathotypes had virulence to Thatcher wheat lines with leaf rust resistance genes Lr23 and Lr42. However, Lr24, LrB, Lr10, Lr14b and Lr20 genes exhibited susceptibility response i.e. (HITs 3 4) with all tested pathotypes. Based on virulence, ten virulence phenotypes (MSCTNS, RTSTNS, RKTRGS, PNDQDS, JDBQGJ, MDPSDS, RTPTPS, MNPSDS, MJLTGS and MSPTDS) were identified among the ten isolates, designated with six-letter code. Two phenotypes RTSTNS RTPTPS exhibited broad spectrum, both were virulent to nineteen resistance genes of leaf rust while pathotype JDBQGJ had narrow spectrum as compared to all other tested, with virulence to just eight resistance genes of leaf rust. Among the locations virulence variability of leaf rust was also recorded. Most of identified races were virulent to more than one of leaf rust resistance genes. Resistance genes (Lr42 and Lr23) identified as effective can be exploited to achieve leaf rust resistance in wheat. Further, the study provides virulence profile of the area may help to manage the leaf rust pathogen.

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