SUPPLEMENTAL DATA
Supplemental Fig. S1 Plastid RNA editing profiles insiliques (Sil),flowers (F),stems
(St), leaves (L),androots (R)from 6-week-old Arabidopsis. Bulk sequencing of the
RT-PCR products containing the RNA editing sites are shown as sequence
chromatograms. Arrows indicate the editing sites.
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Supplemental Fig. S2 RT-PCR analysis of total RNA extracted from roots with
primers used for ndhB-1255 and ndhD-2. PCR products were not detected in reactions
without reverse transcriptase (- RT), indicating that contamination of genomic DNA
was negligible in the analysis of ndhB-1255 and ndhD-2 editing in roots.
Supplemental Fig. S3 RNA gel blot analysis of plastid genes in different organs from
6-week-old Arabidopsis. R, roots; L, leaves; St, stems; F, flowers; Si, siliques.
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Supplemental Fig. S4 Arabidopsis plastid RNA editing profiles were similar in 3day-old green (G) and etiolated (E) seedlings. Bulk sequencing of the RT-PCR
products containing the RNA editing sites are shown as sequence chromatograms.
Arrows indicate the editing sites.
3
Supplemental Fig. S5TheArabidopsis ispG null mutant is albino lethal.(A) The IspG
gene encodes HMBPP synthase (HDS) of the nonmevalonate pathway for IPP and
DMAPP biosynthesis in the plastid, which are precursors of isoprenoids including
carotenoids and chlorophylls. (B) Schematic diagram of the IspG gene and the TDNA insertion site of the ispG mutant (SALK_017595). (C) Progeny of a green
heterozygous ispG mutant segregate green and albino seedlings in a ratio of ~3:1. (D)
RNA gel blot analysis revealed that the IspG transcripts were not detectable in the
ispG albino seedlings. (E) The ispG mutant chloroplast contained large vesicles and
lipid droplets.
4
Supplemental Fig. S6 Plastid RNA editing profilesin wild type (WT),and cia5-2,
ispF, and ispG albino mutants. Bulk sequencing of the RT-PCR products containing
the RNA editing sites are shown as sequence chromatograms. Arrows indicate the
editing sites.
5
Supplemental Fig. S7 Plastid RNA editing profilesin green (G),lincomycin (Lin)and norflurazon (NF)-treated seedlings. Bulk sequencing of the RT-PCR products
containing the RNA editing sites are shown as sequence chromatograms. Arrows
indicate the editing sites.
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Supplemental Fig. S8 Editing of ndhB-467, ndhB-586, and ndhB-746 on spliced and
unsplicedndhBtranscripts.Bulk sequencing results of the RT-PCR products containing
the RNA editing sites are shown as sequence chromatograms. Arrows indicate the
editing sites. These editing sites were sequenced from spliced and unspliced RT-PCR
products purified from the same reaction and amplified by primers located on
different exons. In NF-treated seedlings, both spliced and unsplicedndhB transcripts
were amplified. The editing of ndhB-467, ndhB-586, and ndhB-746 was more affected
in unspliced transcripts. In Lin-treated seedlings, only unsplicedndhB transcripts were
amplified.
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Supplemental Fig. S9 RNA gel blot analysis of Arabidopsis plastid genes in cia5-2,
ispF, and ispG albino mutants. W, wild type; C, cia5-2; F, ispF; G,ispG.
Supplemental Fig. S10 RNA gel blot analysis of Arabidopsis plastid genes in green
(G),lincomycin (L)- and norflurazon (N)-treated seedlings.
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Supplemental Table S1 Primers used for making DIG-labeled probes and probe
locations
Gene
Primer sequence 5'→3'
Probe location
accD
atpF
clpP
matK
ndhB
ndhD
ndhF
ndhG
petL
psbE
psbF
psbZ
rpl23
rpoA
rpoB
rpoC1
GAAAGCGGAAAGATTCGTGA / ATGGAGCTGAAATAACTCGC
TAGATAACCGAAAGCAGAGG / GTTATTTCTTTCATCGTACC
ATGATCCATCAACCCGCTAG / TATTGAACCGCTACAAGATC
CGGCAATCCTAGGGTTGCTC / AAATACCAAATTCGCACTCG
ATGGCTATAACAGAGTTTCT / CGCAATAATCGGGTTCATTG
CATGGATTTATTGGCGCTGC / GATTCGACCTTGTCACTTGC
GTGGCTCCTTCTATTCACTC / GAAGAGATGCGACTTCCACC
GGATTTGCCTGGACCAATAC / TGACGAGCCACAGAAATTGC
ATGCCTACTATAACTAGTTA / TCAAATAAGTCGTATTTTGC
TCTGGAAGCACAGGAGAACG / TACTAAATTCATCGAGTTGT
ATGACTATAGATAGGACCTA / TTATCGTTGGATGAACTGCA
GACTATTGCTTTCCAATTGG / AAGAGATAAGAGAATTAAGG
GGATGGAATCAAATATGCAG / TAAGTTCTTTTCTTTCTAAG
CGGACACTACAGTGGAAGTG / ATGAATACAGCATCGATAGG
ATGCCTTATTTGCAAGACGG / GATCGTAGTTCTCGAACAAG
TTTTCTTTTGCTAGGCCCATAA / TTCGCAAATCTAAATCGGCT
rps 12
ATGCCAACCATTAAACAACT / ATACACCCGAGTACATGTTC
rps 14
ATGGCAAAGAAAAGTTTGAT / TTACCAGCTTGATCTTGTTG
Exon 1
Exon 2
Exon 3
Exon 1
Exon 1+Exon 2
Exon 1
Exon 1
Exon 1
Exon 1 (orf31)
Exon 1
Exon 1
Exon 1
Exon 1
Exon 1
Exon 1
Exon 2
Exon 1
(ATCG00065)
Exon 1
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Supplemental Table S2 Primers used for RT-PCR and analysis of plastid RNA editing
Editing site
Primer sequence 5'-->3'
accD-794
accD-1568
atpF-92
clpP-559
matK-640
TGAAAGCGGAAAGATTCGTG/ATGGAGCTGAAATAACTCGC
TGGATGCAATGGTACCACGTA/AGAATCTGATCTAACAACAGGGAA
CCGATTCTTTCGTTTACTTG/AGGGTTCCTATAGCTCCTTG
ATGATCCATCAACCCGCTAG/TATTGAACCGCTACAAGATC
CGTTACCGGGTAAAAGATGC/AGCGGCGTATCCTTTGTTGC
TTCCGTAAATATTCCATTCCAAA/
ndhB-149
AAACATTCCTCCTAGAGTAGCTGTTAAT
ndhB-467
CATTGAATGTACAGAAATGGCTATAA/TCCGATTTGACCTATGGACG
ndhB-586
CATTGAATGTACAGAAATGGCTATAA/TCCGATTTGACCTATGGACG
ndhB-746
CATTGAATGTACAGAAATGGCTATAA/TCCGATTTGACCTATGGACG
ndhB-830
GGACTCCTGACGTATACGAA/TCCGATTTGACCTATGGACG
ndhB-836
GGACTCCTGACGTATACGAA/TCCGATTTGACCTATGGACG
ndhB-872
GGACTCCTGACGTATACGAA/TCCGATTTGACCTATGGACG
ndhB-1255 GTCGTTGCTTTTCTTTCTG/CTAAAAAGGGTATCCTGAGC
ndhB-1481 GTCGTTGCTTTTCTTTCTG/CTAAAAAGGGTATCCTGAGC
ndhD-2
GACTGTGTTGGTTGTAAGAGATGTG/TCGATCCATTTATAATCTTCGGA
ndhD-383
GGACCTGGTGTATCTTGTC/TAGCCGAATACAGACGTTTC
ndhD-674
TATGGTTCGAACGAACCAAC/CATAACTCGTTCATGCCAAG
ndhD-878
TATGGTTCGAACGAACCAAC/CATAACTCGTTCATGCCAAG
ndhD-887
TATGGTTCGAACGAACCAAC/CATAACTCGTTCATGCCAAG
ndhF-290
AAAACCTTCGCCGCATGTGG/GCATTCGCTGCAATAGGTCG
ndhG-50
ATAATGGATTTGCCTGGACC/CTTATTAAATCTTGCTCTAGAATCTGGTT
petL-5
AGGGAAGTACTTTAAGAATC/ATTAGACCTAAGACGATTCC
psbE-214
ACAGGAGAACGTTCTTTTGC/CGTTGGATGAACTGCATTGC
psbF-77
ACAGGAGAACGTTCTTTTGC/CGTTGGATGAACTGCATTGC
AGAACATAGCCCTATGAGTTAATACGA/
psbZ-50
GATAAGAGAATTAAGGATACCCACCA
rpl23-89
AATTCCTACTGGATGCACGC/AAGAGGTGGAATAGAATAACCCG
rpoA-200
CGGACACTACAGTGGAAGTG/ATGAATACAGCATCGATAGG
rpoB-338
TATCGGTTTATTGATCAGGG/GCAGCTGCTAACACATCTCG
rpoB-551
TATCGGTTTATTGATCAGGG/GCAGCTGCTAACACATCTCG
rpoB-2432
AACACCTCAAGTGGCGAAAG/GTCCTACATTCATGCGTGAG
rpoC1-497
TTTTCTTTTGCTAGGCCCATAA/TTCGCAAATCTAAATCGGCT
rps12-intron CAAGACAGCCAATCCGAAAC/CTTGTACAATTCACATTCTTTGGC
rps14-50
TTCTTAATTTCCTGGCGTGG/TCCATTTCTCACTTAGCGACG
rps14-149
TCATTTGATTCGTCGATCCTC/TACCAGCTTGATCTTGTTGC
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Supplemental materials and methods
The proteins used for amino acid sequence alignments can be found in the
GenBankdatabase under the following accession numbers. Nicotianatabacum: accD,
NP_054508; atpF, NP_054482; clpP, NP_054525; matK, NP_054478; ndhB,
NP_054547; ndhD, NP_054557; ndhF, NP_054554; ndhG, NP_054560; petL,
NP_054519; psbE, NP_054517; psbF, NP_054516; psbZ, NP_054494; rpl23,
NP_054541; rpoA, NP_054532; rpoB, NP_054488; rpoC1, NP_054487; rps14,
NP_054495. Oryza sativa: atpF, NP_039379; clpP, NP_039410; matK, NP_039361;
ndhB, NP_039432; ndhD, NP_039444; ndhF, NP_039441; ndhG, NP_039447; petL,
NP_039404; psbE, NP_039403; psbF, NP_039402; psbZ, NP_039368; rpl23,
NP_039392; rpoA, NP_039417; rpoB, NP_039373; rpoC1, NP_039374; rps14,
NP_039381. Zea mays: atpF, NP_ 043021; clpP, NP_043048; matK, NP_043005;
ndhB, NP_043076; ndhD, NP_043087; ndhF, NP_043084; ndhG, NP_043090; petL,
NP_043042; psbE, NP_043041; psbF, NP_043040; psbZ, NP_043011; rpl23,
NP_043068; rpoA, NP_043055; rpoB, NP_043015; rpoC1, NP_043016; rps14,
NP_043023. Marchantiapolymorpha: accD, NP_039307; atpF, NP_039281; clpP,
NP_039323; ndhB, NP_039271; ndhD, NP_039348; ndhF, NP_039343; ndhG,
NP_039351; psbE, NP_039315; psbF, NP_039314; psbZ, NP_039296; rpl23,
NP_039342; rpoA, NP_039331; rpoB, NP_039275; rpoC1, NP_039276; rps14,
NP_039297. Physcomitrella patens: accD, NP_904193; atpF, NP_904217; clpP,
NP_904176; matK, NP_904210; ndhB, NP_904166; ndhD, NP_904238; ndhF,
NP_904235; ndhG, NP_904241; petL, NP_904182; psbE, NP_904183; psbF,
NP_904184; psbZ, NP_904205; rpl23, NP_904234; rpoB, NP_904223; rpoC1,
NP_904222; rps14, NP_904204. Chlamydomonasreinhardtii: atpF, NP_958410; clpP,
NP_958364; petL, NP_958424; psbE, NP_958396; psbF, NP_958399; psbZ,
NP_958383; rpl23, NP_958368; rpoA, NP_958389; rps14, NP_958381.
Synechocystissp. PCC 6803: atpF, NP_440057; clpP, NP_441889; ndhB, NP_440041;
ndhD, NP_441967; ndhF, NP_442460; ndhG, NP_442812; psbE, NP_440412; psbF,
NP_440413; psbZ, NP_440888; rpl23, NP_440668; rpoA, NP_440645; rpoB,
NP_440685; rpoC1, NP_441586.
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