5.4.99.8 - 5.4.99.8 cloning of full-length gene WsCAS by homology-based PCR method, DNA and amino acid sequence determination and analysis, sequence comparisons, transformation of overexpressing and RNAi constructs into Withania somnifera plants via Agrobacterium strain GV3101 transformation method, quantitative RT-PCR enzyme expression analysis 5.4.99.8 DNA and amino acid sequence determination and analysis, sequence comparisons and phylogenetic analysis, cloning and expression in Escherichia coli strain D5alpha, functional expression of GST-tagged enzyme in Schizosaccharomyces pombe 5.4.99.8 expressed in Escherichia coli and in the yeast mutant GIL77 5.4.99.8 expressed in Saccharomyces cerevisiae mutant GIL77 cells 5.4.99.8 expression in DH5-alpha Escherichia coli competent cells 5.4.99.8 expression in Escherichia coli 5.4.99.8 expression in Sacchaormyces cerevisiae 5.4.99.8 expression in Saccharomyces cerevisiae 5.4.99.8 expression in yeast 5.4.99.8 full-length cDNAs of SgCAS is cloned by a rapid amplification of cDNA-ends with polymerase chain reaction approach. Transient expression in Nicotiana benthamiana 5.4.99.8 gene CAS, recombinant expression in TBY-2 cells, usage of Agrobacterium tumefaciens strain LBA4404 hyper-virulent strain and transformation into BY-2 cell suspensions via co-culture, quantitative reverse transcriptase-mediated real-time PCR enzyme expression analysis. After 6 days of treatment, the expression of NtCAS1 increases 2.5fold in the BY-2 cells 5.4.99.8 gene CAS1, sequence comparisons and phylogenetic tree 5.4.99.8 gene CAS1, sequence comparisons and phylogenetic tree, recombinant expression in the Saccharomyces cerevisiae mutant erg7 that lacks an endogenous lanosterol synthase. Upon galactose-induced expression of NtCAS1, yeast cells grown in the presence of exogenous ergosterol display a sterol profile that includes cycloartenol and four other sterols in addition to ergosterol. The major sterol is 24-methylene pollinastanol that accounts for 53% of the total and represents the most probable pathway end-product generated by the yeast steroidogenic machinery, apparently being versatile enough to accept cycloartenol as a substrate 5.4.99.8 gene is present in single copy. Expression of enzyme in Saccharomyces cerevisiae 5.4.99.8 gene LdCAS, DNA and amino acid sequence determination and analysis, sequence comparisons and phylogenetic analysis, functional recombinant expression in a sterol-engineered Saccharomyces cerevisiae strains PA14, TM097, and TM122, that are derived from strain TM1 in the S288c / BY4742 background. Yeast strain PA14 is derived from strain TM1 by knocking out the TRP1 gene using CRISPR/Cas9 5.4.99.8 gene SgCAS, DNA and amino acid sequence determination and analysis, sequence comparisons and phylogenetic analysis, gene SgCAS belongs to the cycloartenol synthase cluster, quantitative real-time PCR enzyme expression analysis, transcriptome analysis, recombinant expression in Escherichia coli strain BL21(DE3), transient expression of GFP-tagged SgCAS in Nicotiana benthsmiana lower epidermal cells 5.4.99.8 isozyme CAS1, RNA-sequencing analysis from seedling genome, cloning from cDNA library, DNA and amino acid sequence determination and analysis, isozyme sequence comparison, phylogenetic tree of OSCs, quantitative RT-PCR enzyme expression analysis, functional recombinant expression in Saccharomyces cerevisiae, GC-MS analysis of cycloartenol in yeast 5.4.99.8 isozyme CAS2, RNA-sequencing analysis from seedling genome, cloning from cDNA library, DNA and amino acid sequence determination and analysis, isozyme sequence comparison, phylogenetic tree of OSCs, quantitative RT-PCR enzyme expression analysis, functional recombinant expression in Saccharomyces cerevisiae, GC-MS analysis of cycloartenol in yeast 5.4.99.8 mutants are expressed in Saccharomyces cerevisiae strain LHY4 5.4.99.8 on the base of callus cells, enzyme CAS recombinant expression in Panax notoginseng cells using Agrobacterium tumefaciens strain EHA105 for transfection, coexpression with farnesyl diphosphate synthase (FPS). Plasmid pHellsgate-CAS is transformed into the wild-type cells, and pCAMBIA1300S-FPS is then introduced into the pHellsgate-CAS transformed cells. Quantitative real-time RT-PCR enzyme expression analysis 5.4.99.8 sequence comparisons of lanosterol synthases and cycloartenol synthases from several organisms