Dataset: HRR 18S full-length sequence BLAST results

Name: Results of 18S sequencing of full-length 18S rDNA for metabarcoding samples collected during R/V Point Sur cruise PS18-09 in the Western Gulf of Mexico in September 2017
Published: 2024-03-14
Keywords: oceans

Cite This Dataset

DOI:10.26008/1912/bco-dmo.922101.1

Spatial Extent: N:29.0649 E:-94.9 S:27.8358 W:-96.9874

Gulf of Mexico, Texas coast

Temporal Extent: 2017-09-23

Project:

RAPID: Hurricane Impact on Phytoplankton Community Dynamics and Metabolic Response (HRR)

Principal Investigator:

Lisa Campbell (Texas A&M University, TAMU)

Co-Principal Investigator:

Darren W. Henrichs (Texas A&M University, TAMU)

Contact:

Chetan Gaonkar (Texas A&M University, TAMU)

BCO-DMO Data Manager:

Shannon Rauch (Woods Hole Oceanographic Institution, WHOI BCO-DMO)

Version:

Version Date:

2024-03-14

Restricted:

Validated:

Yes

Current State:

Final no updates expected

Results of 18S sequencing of full-length 18S rDNA for metabarcoding samples collected during R/V Point Sur cruise PS18-09 in the Western Gulf of Mexico in September 2017

Abstract:

This dataset includes results of 18S sequencing of full-length 18S rDNA for metabarcoding samples collected from surface depth at stations 06, 11, 16, 21, SS, and GI during R/V Pt. Sur cruise PS18-09 Leg 1 using Nanopore minION.

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Archival Copy

Version Date	Archive	Persistent Identifier (PID)	Date PID Issued
2024-03-14	Marine Biological Laboratory/Woods Hole Oceanographic Institution Library (MBLWHOI DLA)	10.26008/1912/bco-dmo.922101.1	2024-03-15

Methods & Sampling

On September 23, 2017, on HRR cruise Leg 1 (R/V Point Sur PS18-09), samples were collected at 6 stations (S06, S11, S16, S21, SS, and GI) from the surface depth bottle in a CTD rosette. Triplicate 500-1000 milliliter (mL) samples were filtered and immediately fixed in RNALater. Triplicate samples from each station/depth were extracted with AllPrep DNA/RNA MiniKit (Qiagen, USA) following the manufacturer's instructions. DNA concentration and quality were evaluated using a Nanodrop spectrophotometer (Thermo Fisher Scientific Inc, USA). All the samples extracted for DNA were normalized to 5 nanograms per microliter (ng/µL) concentration for the amplicon library construction.

The full-length 18S rDNA region was amplified using 18S primers (Medlin et al. 1988; Edgar & Theriot 2004). Library construction and amplicon sequencing were performed at the Campbell Lab, Texas A&M University using Oxford Nanopore minION. The output sequencing results as fasta files were deposited in NCBI's GenBank under the project number PRJNA592369.

Data Processing Description

Fastq files for all six station samples were quality checked using FastQC. Resulting sequences were taxonomically annotated using BLAST search against PR2 database v5.0.0. The BLAST analysis used an assignment approach with similarity was ≥ 90% and query coverage was ≥ 1000 bp against the reference sequence. Any sequence that did not compile with this criterion was not used in this study. The following thresholds for identity with BLAST results were used for taxonomic assignment clustering: genus (95%), family (93%), class (92%), and order (90%). Archaeal and Bacterial sequences were removed from these results.

Additional tools used are:
Oxford Nanopore Technologies. Guppy protocol. Available at: https://community.nanoporetech.com/docs/prepare/library_prep_protocols/Guppy-protocol/v/gpb_2003_v1_revax_14dec2018/guppy-software-overview [Accessed 21 Aug 2023].

BCO-DMO Processing

- Saved the location table (station number, lat, lon, etc.) submitted as part of the dataset metadata in a separate spreadsheet.
- Imported the 6 original CSV files (provided in zip "HRR_18S_BLAST_results.zip") into the BCO-DMO system.
- Concatenated the 6 CSV files into a single data table.
- Joined the columns from the location table to the primary data table by matching on Station number.
- Saved final file as "922101_v1_hrr_18s_blast_results.csv".

Data Files

File
922101_v1_hrr_18s_blast_results.csv (Comma Separated Values (.csv), 117.16 MB) MD5:6ec461e1f2a8df4ba226050af4974d59 Primary data file for dataset ID 922101, version 1

More Information about this dataset

Funding Sources

Funding Source	Award Number
NSF Division of Ocean Sciences	OCE-1760620

Deployments

Deployment	Synonyms	Start Date	Platform	Investigator
PS1809	HRR PS18-09	2017-09-22	R/V Point Sur	Lisa Campbell (Chief Scientist) Steven DiMarco (Chief Scientist)	data info

Instruments

Automated DNA Sequencer

Supplied Name: Oxford Nanopore minION Mk1C

Supplied Description:

Library construction and amplicon sequencing was performed at Campbell Lab, Texas A&M University using Oxford Nanopore minION.

Instrument Type

Generic Name: Automated DNA Sequencer

Acronym: Automated Sequencer

Generic Description:

General term for a laboratory instrument used for deciphering the order of bases in a strand of DNA. Sanger sequencers detect fluorescence from different dyes that are used to identify the A, C, G, and T extension reactions. Contemporary or Pyrosequencer methods are based on detecting the activity of DNA polymerase (a DNA synthesizing enzyme) with another chemoluminescent enzyme. Essentially, the method allows sequencing of a single strand of DNA by synthesizing the complementary strand along it, one base pair at a time, and detecting which base was actually added at each step.

CTD - profiler

Supplied Name: CTD rosette bottle

Supplied Description:

Samples were collected from the surface depth bottle in a CTD rosette.

Instrument Type

Generic Name: CTD - profiler

Community Identifier: http://vocab.nerc.ac.uk/collection/L05/current/130/

Generic Description:

The Conductivity, Temperature, Depth (CTD) unit is an integrated instrument package designed to measure the conductivity, temperature, and pressure (depth) of the water column. The instrument is lowered via cable through the water column. It permits scientists to observe the physical properties in real-time via a conducting cable, which is typically connected to a CTD to a deck unit and computer on a ship. The CTD is often configured with additional optional sensors including fluorometers, transmissometers and/or radiometers. It is often combined with a Rosette of water sampling bottles (e.g. Niskin, GO-FLO) for collecting discrete water samples during the cast.

This term applies to profiling CTDs. For fixed CTDs, see https://www.bco-dmo.org/instrument/869934.

Spectrophotometer

Supplied Name: Nanodrop spectrophotometer

Supplied Description:

DNA concentration and quality were evaluated using a Nanodrop spectrophotometer (Thermo Fisher Scientific Inc, USA).

Instrument Type

Generic Name: Spectrophotometer

Community Identifier: http://vocab.nerc.ac.uk/collection/L05/current/LAB20/

Generic Description:

An instrument used to measure the relative absorption of electromagnetic radiation of different wavelengths in the near infra-red, visible and ultraviolet wavebands by samples.

Parameters

Date and time formats are described in python strftime() syntax.

Supplied Name	Supplied description	Supplied Units	Standard Name
Station	sampling station number	unitless	station
Leg	cruise leg	unitless	leg
Latitude	latitude of sampling station; positive values = North	decimal degrees	lat
Longitude	longitude of sampling station; negative values = West	decimal degrees	lon
Sequence_ID	OTU sequence generated from the HRR data	unitless	sequence
Taxonomy	taxonomic annotation of the O.T.U.	unitless	taxon
Similarity	percentage of identical matches	percent	sample_descrip
length	alignment length (sequence overlap)	base pairs (bp)	length
mismatches	number of mismatches	base pairs (bp)	sample_descrip
gaps	number of gap openings	unitless	sample_descrip
Q_start	start of alignment in query	base pairs (bp)	sample_descrip
Q_end	end of alignment in query	base pairs (bp)	sample_descrip
R_start	start of alignment in reference sequence	base pairs (bp)	sample_descrip
R_end	end of alignment in reference sequence	base pairs (bp)	sample_descrip
e_value	number of expected hits of similar quality	unitless	sample_descrip
score	Bit-score	unitless	sample_descrip
file_name_orig	original name of the csv file for this station+leg	unitless	file_name