Next-Generation Sequencing of Haematococcus lacustris Reveals an Extremely Large 1.35-Megabase Chloroplast Genome

Bauman, Nicholas; Akella, Srividya; Hann, Elizabeth; Morey, Robert; Schwartz, Ariel; Brown, Rob; Richardson, Toby H.

doi:10.1128/genomea.00181-18

Cited by 32 publications

(31 citation statements)

References 16 publications

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“…In terms of organellar DNA size, the current record holder is the parasitic plant Silene conica , which has a mitochondrial genome of 11.3 Mbp, most of which has been acquired by horizontal gene transfer . The largest plastid genomes sequenced thus far are the 1.13 Mbp genome of the red alga Corynoplastis japonica and that of the green alga Haematococcus lacustris , whose genome is 1.35 Mbp in size . To our knowledge, total non‐nuclear DNA content has not been estimated for any of these organisms.…”

Section: Resultsmentioning

confidence: 99%

Massive mitochondrial DNA content in diplonemid and kinetoplastid protists

et al. 2018

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SummaryThe mitochondrial DNA of diplonemid and kinetoplastid protists is known for its suite of bizarre features, including the presence of concatenated circular molecules, extensive trans‐splicing and various forms of RNA editing. Here we report on the existence of another remarkable characteristic: hyper‐inflated DNA content. We estimated the total amount of mitochondrial DNA in four kinetoplastid species (Trypanosoma brucei, Trypanoplasma borreli, Cryptobia helicis, and Perkinsela sp.) and the diplonemid Diplonema papillatum. Staining with 4′,6‐diamidino‐2‐phenylindole and RedDot1 followed by color deconvolution and quantification revealed massive inflation in the total amount of DNA in their organelles. This was further confirmed by electron microscopy. The most extreme case is the ∼260 Mbp of DNA in the mitochondrion of Diplonema, which greatly exceeds that in its nucleus; this is, to our knowledge, the largest amount of DNA described in any organelle. Perkinsela sp. has a total mitochondrial DNA content ~6.6× greater than its nuclear genome. This mass of DNA occupies most of the volume of the Perkinsela cell, despite the fact that it contains only six protein‐coding genes. Why so much DNA? We propose that these bloated mitochondrial DNAs accumulated by a ratchet‐like process. Despite their excessive nature, the synthesis and maintenance of these mtDNAs must incur a relatively low cost, considering that diplonemids are one of the most ubiquitous and speciose protist groups in the ocean. © 2018 The Authors. IUBMB Life published by Wiley Periodicals, Inc. on behalf of International Union of Biochemistry and Molecular Biology., 70(12):1267–1274, 2018

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Section: Resultsmentioning

confidence: 99%

Massive mitochondrial DNA content in diplonemid and kinetoplastid protists

et al. 2018

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“…For instance, the chloroplast genome of the green alga Haematococcus lacustris , which is currently the largest on record (1.35 Mb), was recently published in the journal Genome Announcements . Sadly, the GenBank entry accompanying this paper lacked even the most straightforward annotations, such as ribosomal RNAs, and contained a number of mislabelled genes.…”

Section: Organelle Genomics a Victim Of Its Own Successmentioning

confidence: 99%

Revisiting published genomes with fresh eyes and new data

Smith

2019

EMBO Reports

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“…By contrast, cloning the 5’ and 3’ flanking regions of genes in H. pluvialis is still based on genomic walking or screening of genomic libraries. In 2018, the chloroplast genome of Haematococcus lacustrisis was deposited in the Genebank using PacBio long-read sequencing technology [21]; however, its nuclear genome is still unreported. This situation, limits the quick access to and application of transcriptional regulatory sequence in the H.pluvialis nuclear genome.…”

Section: Discussionmentioning

confidence: 99%

Biolistic transformation of Haematococcus pluvialis with constructs based on the flanking sequences of its endogenous alpha tubulin gene

Yuan

Zhang

et al. 2018

Preprint

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18The complete sequence information of the alpha tubulin (tub) genes was obtained from 19 both Haematococcus pluvialis NIES144 and SCCAP K0084. , Putative transcriptional 20 elements and polyadenylation signals were identified respectively in their 5' and 3' 21 flanking regions. Three selection cassettes of tub/aadA, tub/hyr and tub/ble with 3 22 different antibiotic-resistant genes fused between the 5' and 3' flanking sequences of the 23 tub gene were constructed and utilized for biolistic transformation of H.pluvialis. 24 Antibiotic resistant transformants were obtained in the bombardments with the tub/aadA 25 cassette in 2 strains. It was found that, the foreign tub/aadA DNA could be completely 26 transferred and inherited in their genome through non-homologous recombination. 27 Moreover, transcripts of the insert and spectinomycin resistance were identified. 28 Transformation efficiencies up to 3×10 -5 per μg DNA could be obtained in H.pluvialis 29 NIES144 or SCCAP K0084 through utilization of a culture with a high percentage of 30 flagellate cells and by optimizing bombarding protocol. The presented selection marker 31 and optimized transforming procedures in this report should strengthen the platform 32 technology for genetical manipulation and modification of H.pluvialis. 33 34 35 36 37 38 39 3 40 41 Astaxanthin (3, 3'-dihydroxy-ß-carotene-4, 4'-dione) is the strongest antioxidant in the 42 nature, and has been widely used as an additive in health care products and cosmetics [1]. 43 Haematococcus pluvialis has attracted great attention as its richest biological source. 44 Currently, H.pluvialis cultivation has been commercialized to produce astaxanthin 45 powder, which can be utilized as feedstocks or as additives in aquacultural, 46 pharmaceutical and nutraceutical products [2]. H.pluvialis has been considered as an 47 organism for the production of recombinant pharmaceutical proteins[3]. 48 Researches on H.pluvialis have taken place over several decades, mostly focused on 49 cell differentiation [4], astaxanthin metabolism[5], cultivation methods [6] and 50 astaxanthin extraction technology[7]. Continuous efforts were made to improve its 51 cellular growth and the level of astaxanthin accumulation through traditional mutagenesis 52 of H.pluvialis [8]; however there are no reports of phenotypic modification through 53 genetic engineering methods. To date, only 4 researchers have reported stable genetic 54 transformation of H.pluvialis. Steinbrenner & Gerhard Sandmann in 2006 [9] 55 transformed H.pluvialis with a mutated phytoene desaturase (pds) gene conferring 56 resistance to norflurazon, using a biolistic method. Sharon-Gojman et al. in 2015[10] 57 optimized this vector system by shortening the promoter sequence from 2000 base pairs 58 (bp) to 1000 bp. And an agrobacterium-mediated method using a plant-general plasmid 59 pCAMBIA1301 could transform H.pluvialis, with a foreign hptII gene been identified in 60 the transformants [11]. Successful chloroplast transformation was also achieved thr...

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Next-Generation Sequencing of Haematococcus lacustris Reveals an Extremely Large 1.35-Megabase Chloroplast Genome

Cited by 32 publications

References 16 publications

Massive mitochondrial DNA content in diplonemid and kinetoplastid protists

Massive mitochondrial DNA content in diplonemid and kinetoplastid protists

Revisiting published genomes with fresh eyes and new data

Biolistic transformation of Haematococcus pluvialis with constructs based on the flanking sequences of its endogenous alpha tubulin gene

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