Differential Expression of the Transcripts of Spartina alterniflora Loisel (Smooth Cordgrass) Induced in Response to Petroleum Hydrocarbon

Abstract

Petroleum hydrocarbons (PHC) in soil are potentially toxic to plants and exert negative effect on the environment and human health. To understand the effect of PHC on the gene expression profile of a wetland plant Spartina alterniflora in the coastal Louisiana, plants were subject up to 40% PHC under greenhouse conditions. The plants exposed to PHC showed 21% reduction of leaf total chlorophyll after 2 weeks of stress. Using 20 annealing control primers, 28 differentially expressing genes (DEGs) were identified in leaf and root tissues of S. alterniflora in response to PHC stress. Eleven of these 28 DEGs had role in either molecular function (chlorophyll a-b binding protein, HSP70, NADH, RAN1-binding protein, and RNA-binding protein), biological processes (cell wall protein, nucelosome/chromatin assembly factor) or cellular function (30 S ribosomal protein). This indicated that genes in different regulatory pathways of S. alterniflora were involved in response to PHC. All DEGs showed reduced transcript accumulation in root under oil stress, whereas they showed up- or down-regulation in their transcript abundance in leaf depending on the concentration of the PHC. The genes identified through this study could be used in the genetic screen of S. alterniflora for resistance to PHC.

Identification and Expression of Skinning Injury-responsive Genes in Sweetpotato

“…First-strand cDNA synthesis was performed using a Gene-Fishing TM DEG premix kit (Seegene, Rockville, MD) as previously described (Ramanarao et al, 2011). Briefly, 3 mg of RNA representing different time points (0, 2, 4, 8, and 12 h) was reverse-transcribed to first-strand cDNA at 42°C for 90 min in a final reaction volume of 20 mL containing 1· reaction buffer, 2 mM dNTP mix, 20 U RNase inhibitor (Promega, Madison, WI), 2 mL of 10 mM (dT)15-ACP1 (Seegene), and 1 mL Moloney murine leukemia virus transcriptase [200 U/mL (Promega)].…”

“…Second-strand cDNA synthesis and subsequent PCR amplification were performed in a single tube as previously described (Ramanarao et al, 2011). Second-strand cDNA was synthesized by first-stage PCR followed by a second-stage PCR in a final reaction volume of 20 mL containing 50 ng of the diluted first-strand cDNA, 1· SeeAmp ACP TM master mix (Seegene), 1 mL of 10 mM dT-ACP2, and 2 mL of 5 mM arbitrary ACPs [Seegene (Ramanarao et al, 2011)]. The tube containing the reaction mixture was placed in a preheated (94°C) thermal cycler.…”

“…Two microliters of the first-strand cDNA was used for PCR analysis using DEG-specific primers (Table 1) designed using Primer 3.0 (Rozen and Skaletsky, 2000). The PCR conditions were the same (annealing at 55°C for 35 cycles) as described earlier (Ramanarao et al, 2011). 212 QUANTITATIVE REVERSE TRANSCRIPTION-PCR (QRT-PCR) OF DEGS.…”

“…Because of the high annealing specificity during PCR, the application of the ACP to DEG identification generates reproducible, accurate, and long (100 bp to 2 kb) PCR products that are detectable on agarose gels (Kim et al, 2004). This technique has been used to identify abundant, transient, and rarely expressed candidate genes induced in response to Aspergillus flavus infection in cotton [Gossypium hirsutum (Lee et al, 2012)] and petroleum hydrocarbon exposure in Spartina alterniflora (Ramanarao et al, 2011). The objective in this study was to directly identify critical physiological processes involved in the healing process by examining gene expression changes after intentional skinning.…”

Transcriptome response of the foundation plant Spartina alterniflora to the Deepwater Horizon oil spill

“…Spartina alterniflora also shows remarkable resilience to a variety of stressors (Baisakh & Subudhi, 2009;Baisakh, Subudhi, & Varadwaj, 2008;Pennings & Bertness, 2001;Silliman et al, 2012). Oil-affected populations showed up to 100% recovery within seven months of the DWH spill, despite the immediate effects of reduced carbon fixation and transpiration (Lin & Mendelssohn, 2012;Lin et al, 2016;RamanaRao, Weindorf, Breitenbeck, & Baisakh, 2012;Silliman et al, 2012), and evidence for genetic divergence of oil-exposed populations from nearby uncontaminated populations (Robertson, Schrey, Shayter, Moss, & Richards, 2017). Prior studies in S. alterniflora under controlled conditions have examined expression of candidate genes in response to heat, salt or oil stressors (Baisakh & Subudhi, 2009;Baisakh et al, 2008;RamanaRao et al, 2012), and Bedre, Mangu, Srivastava, Sanchez, and Baisakh (2016) recently characterized the full transcriptome response to salinity.…”