RcbL gene expression/quant

• Page authors: PRIMO
• Responsible curator: Hagen Buck-Wiese

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RuBisCO gene expression (rbcL)
Approach: RT-qPCR or ddPCR of rbcL transcripts or gene copies
Context: in situ, incubation, lab
Spatial scale: point sample
Temporal scale: minutes to hours
Units: transcripts L-1 or relative abundance
Community captured: group specific (phytoplankton)
Co-measurements: 13C- or 14C-based photosynthesis–irradiance curves (Pmax)

Method Overview

The rbcL gene encodes the large subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (RuBisCO), the primary enzyme for carbon fixation in oxygenic photosynthesis. Quantification of rbcL gene copies (from DNA) provides an index of the abundance of photosynthetically competent phytoplankton carrying Form I or Form II RuBisCO, while rbcL transcript abundance (from RNA, via RT-qPCR) reflects transcriptional activity of the gene. When combined with carbon fixation measurements (e.g., P–E curves), rbcL expression can be used to link molecular data to carbon fixation rates.

Degenerate primers targeting conserved rbcL regions are used to survey a broad range of phytoplankton taxa. Group-specific primers (e.g., for diatoms, dinoflagellates, or cyanobacteria) provide more taxonomically resolved information.

Scale of measurement

Each filtered sample provides a point measurement. Transcripts reflect expression at time of sampling, which varies with light history and can change on timescales of minutes to hours under fluctuating irradiance.

Data generated

rbcL transcript concentrations (transcripts L^-1) or gene copies (copies L^-1). When expressed as relative abundance (normalized to a housekeeping gene), transcriptional responses to light, nutrients, or other treatments can be quantified.

Units & currency

Units are transcripts L^-1 or relative abundance. The currency is cDNA (for RNA-based assays) or DNA copies.

Sample size

Typical samples are ~1 L in volume.

Limitations

rbcL transcript abundance does not translate directly to a carbon fixation rate without an empirical coupling factor (typically derived from paired ¹⁴C measurements). Degenerate primers required for broad-spectrum environmental surveys introduce PCR biases that may over- or under-represent certain taxa. A single rbcL copy per genome is assumed, which may not hold for all taxa. Post-transcriptional regulation means transcripts can accumulate without corresponding increases in protein or enzymatic activity.

Example Applications & Protocols

Classic examples

• Pujari et al. (2019) Diversity and spatial distribution of chromophytic phytoplankton in the Bay of Bengal revealed by RuBisCO genes (rbcL)

Recent applications

• Endo et al. (2017) Phytoplankton community responses to iron and CO₂ enrichment in different biogeochemical regions of the Southern Ocean

Common calculations/conversions

• rbcL-based carbon fixation rate requires empirical calibration: GPP = rbcL transcripts × K_rbcL, where K_rbcL is determined from paired ¹⁴C or O₂ measurements.