cDNA-AFLP analysis of gene expression differences between the flower bud and sprout-shoot apical meristem of Angelica sinensis (Oliv.) Diels

Abstract

Angelica sinensis (Oliv.) Diels (Umbelliferae) is a well-known medicinal plant mainly distributed in Gansu Province of China. Its local and global demand is significant because of its food and medicinal applications. However, the early bolting rate of Angelica sinensis (Oliv.) Diels reaches 20%–60%, which seriously affects its food and medicinal qualities. Thus, differences in gene expression between the flower bud and sprout-shoot apical meristem underwent analysis, by means of cDNA-amplified restriction fragment length polymorphism, to better understand the flowering mechanism. 64 primer sets, each of which amplified to 60 transcript-derived fragments (TDFs), were used. Among these TDFs, 26 were expressed specifically in the flower bud. After cloning and sequencing, 32 distinct sequences were obtained from these 26 TDFs, and 25 were found with homologous sequences in databases. Confirmation of differential expression of 13 sequences was obtained by semi-quantitative RT-PCR, their showing higher expression levels in flower buds. These homologous sequences encode transposable elements, pentatricopeptide repeat-containing proteins, DNA-binding transcription factors, zinc finger (B-box type) family proteins, NADP-dependent sorbitol 6-phosphate dehydrogenase (S6PDH), amongst others.

Transcriptome-wide mining of the differentially expressed transcripts for natural variation of floral organ size in Physalis philadelphica

“…Phenotypic variations in general result from alterations in gene activities; therefore, it is likely that the differentially expressed TDFs formed genetic variations causing natural phenotypic variations. Since genome or transcriptome data are not available for Physalis , cDNA-AFLP technology, which is an efficient method of global transcriptional analysis for any species without a reference transcriptome ( Ganeshan et al ., 2011 ; Yu et al ., 2011 ; Yang et al , 2012), was exploited to search for genes causing natural variation in the observed organ size. The 263 cloned differentially expressed TDFs have complicated expression patterns and they encode regulatory factors, protein kinases, protein phosphatase, and various enzymes, which contribute to the complexity of the genetic basis for the different floral phenotypes.…”

Cloning and characterization of the CarbcL gene related to chlorophyll in pepper (Capsicum annuum L.) under fruit shade stress

“…A total of 80 reproducible TDFs were identified with 64 primer combinations tested. All of these TDFs were cloned and sequenced as described previously (Vuylsteke et al, 2007 ; Yu et al, 2011 ; Lin et al, 2014 ). All differentially expressed gene sequences were analyzed using the basic local alignment search tool (BLAST) at the National Center for Biotechnology Information (NCBI) website ( http://www.ncbi.nlm.nih.gov/blast/ ).…”

“…Pre-amplification was performed with the following pre-amplification primers: EcoR I: 5′-GACTGCGTACCAATTCA-3′, Mse I: 5′-GATGAGTCCTGAGTAAC-3′. The resulting products were used as templates for selective amplification (Vuylsteke et al, 2007 ; Yu et al, 2011 ) with selective primers (Supplementary Table 1 ). The PCR products were resolved via electrophoresis on a 4% denaturing polyacrylamide gel, which was then silver-stained and scanned.…”

“…The PCR products were resolved via electrophoresis on a 4% denaturing polyacrylamide gel, which was then silver-stained and scanned. The fragments of interest were excised from the gel and re-amplified using the same primer combinations and PCR conditions used in the selective amplification (Vuylsteke et al, 2007 ; Yu et al, 2011 ). All differentially expressed gene sequences were analyzed using the basic local alignment search tool (BLAST) at the National Center for Biotechnology Information (NCBI) website ( http://www.ncbi.nlm.nih.gov/blast/ ) (Vuylsteke et al, 2007 ; Yu et al, 2011 ).…”