Characterization of the rice RbcS3 promoter and its transit peptide for use in
chloroplast targeted expression
Su-Hyun Park1†, Jin Seo Jeong1†, Yang Do Choi1,2 and Ju-Kon Kim1*
1
Crop Biotechnology Institute, GreenBio Science and Technology, Seoul National University,
Pyeongchang 25354, Korea
2
Department of Agricultural Biotechnology, Seoul National University, Seoul 151-921, Korea
*Corresponding author: Ju-Kon Kim, Crop Biotechnology Institute, GreenBio Science and
Technology, Seoul National University, Pyeongchang 25354, Korea.
Email: jukon@snu.ac.kr. Tel.: +82 33 339 5826. Fax: +82 33 339 5825.
Supplementary Materials
Supplemental Table S1 Median values of relative promoter activity in T2 transgenic 30-dayold leaves and roots
Supplemental Table S2 Primer lists used in this study
Supplemental figure S1 cDNA alignment of the four RbcS genes using CLUSTALW (Larkin
et al. 2007).
Supplemental figure S2 Expression profiles of the four rice RbcS gene in the developmental
stages and tissues.
Supplemental Table S1 Median values of relative promoter activity in T2 transgenic 30-dayold leaves and roots
Constructs
Leaf
Root
RbcS3:TP3:GFP
957
13
RbcS3:GFP
187
175
PGD1:GFP
1618
5863
Supplemental Table S2 Primer lists used in this study
Gene
Specificity
RbcS3
gene
RbcS1
gene
OsCc1
gene
OsUbi1
gene
gfp
gene
RbcS3:TP3 promoter
RbcS3
promoter
TP3
fragment
TP1
fragment
5'5'5'5'5'5'5'5'5'-
Primer Sequence
Forward
Reverse
TATACAGAGGAGACTCGATTGA
-3' 5'- TACATACACAGCTGATGTTGAC
GGCAGGTACTGGACCATGTG
-3' 5'- TTGTCGAAGCCGATGATACG
ACTCTACGGCCAACAAGAAC
-3' 5'- CTCCTGTGGCTTCTTCAACC
ATGGAGCTGCTGCTGTTCTA
-3' 5'- TTCTTCCATGCTGCTCTACC
CAGCACGACTTCTTCAAGTCC
-3' 5'- CTTCAGCTCGATGCGGTTCAC
AAAAAGCAGGCT-GCGAGGTGCTTAGGCTATTG
-3' 5'- AGAAAGCTGGGT-GCATACAGCTGATCCTTCCAC
AAAAAGCAGGCT-GCGAGGTGCTTAGGCTATTG
-3' 5'- AGAAAGCTGGGT-GGTGAGGTGCTAGAGCAGAG
ATGGCGCCCACCGTGATGGC
-3' 5'- ACAGCTGATCCTTCCACCGT
ATGGCCCCCTCCGTGATGGC
-3' 5'- GCACCTGATCCTGCCGCCAT
-3'
-3'
-3'
-3'
-3'
-3'
-3'
-3'
-3'
RbcS4
RbcS1
RbcS2
RbcS3
-----------------------------------------------------------------------------------------------------------------------GCGTGAGCCAGCCACAAACGCCCGCGGATAGGCGCGCGCACGCCGGCCAATCCTACCACATCCCCGGCCTCCGCGGCTCGCGAGCGCCGCTGCCATCCGATCCGCTGAGTTTTGGCTATT
-----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
RbcS4
RbcS1
RbcS2
RbcS3
--------------------GTACTCAAGCCGGAGGC----AGCACACTGCAACTTAAGTTTTTCTATAGCTCCTAGCAAGCTAGCAATGGCTCCCTCGGTGATGGCTTCGTCGGCCACC
TATACGTACCGCGGGAGCCTGTGTGCAGAGCAGTGCATCTCAAGAAGTACTCGAGCAAAGAAGGAGAGAGCTTGGTGAGCTGCAGAGATGGCCCCCTCCGTGATGGCGTCGTCGGCCACC
-------------AACAGCACTGCTACTGGACATACTCTACTACTACTAG-------CCAGTAAGCTAGCTAACTAACTACGTGGCTATGGCCCCCACCGTGATGGCCTCCTCGGCCACC
--------------AAGCCACTGCTGC--AGCGTGACTCTGCTCTAGCAC-------CTCACCAAGCAGAAAGCTAGAGAGCTAGCAATGGCGCCCACCGTGATGGCCTCGTCGGCCACC
*
* ***** *** * ******** ** *********
RbcS4
RbcS1
RbcS2
RbcS3
TCCGTGGCTCCCTTCCAGGGGCTCAAGTCCACTGCGGGCCTCCCGGTGAACCGCCGCTCCAGCAGCTCGAGCTTTGGCAACGTCAGCAATGGCGGAAGGATCAGATGCATGCAGGTGTGG
ACCGTCGCTCCCTTCCAGGGGCTCAAGTCCACCGCCGGCATGCCCGTCGCCCGCCGCTCCGGCAACTCCAGCTTCGGCAACGTCAGCAATGGCGGCAGGATCAGGTGCATGCAGGTGTGG
TCCGTGGCTCCATTCCAAGGGCTCAAGTCCACCGCCGGGCTCCCCGTCAGCCGCCGCTCCACCAACTCGGGCTTCGGCAACGTCAGCAATGGCGGAAGGATCAAGTGCATGCAGGTGTGG
TCCGTTGCTCCCTTCCAGGGGCTCAAATCCACCGCCGGGCTCCCCGTCAGCCGCCGCTCCAACAGCGCCGGCCTCGGCAGCGTCAGCAACGGTGGAAGGATCAGCTGTATGCAGGTGTGG
**** ***** ***** ******** ***** ** ** * ** **
********** ** * * ** * **** ********* ** ** ******* ** ************
RbcS4
RbcS1
RbcS2
RbcS3
CCGATTGAGGGCATTAAGAAGTTCGAGACTCTGTCGTACTTGCCGCCATTGACAGTGGAGGACCTTTTGAAGCAGATCGAGTACCTGCTCCGATCTAAGTGGGTGCCTTGCTTGGAGTTC
CCGATTGAGGGCATCAAGAAGTTCGAGACCCTCTCCTACCTGCCACCGCTCACCGTGGAGGACCTCCTGAAGCAGATCGAGTACCTGCTCCGTTCCAAGTGGGTGCCCTGCCTCGAGTTC
CCAATTGAGGGCATCAAGAAGTTCGAGACCCTATCGTACCTGCCACCACTCACCGTGGAGGATCTTTTGAAGCAGATCGAGTACCTGCTTCGATCCAAGTGGGTGCCTTGCCTCGAGTTC
CCGATTGAAGGCATCAAGAAGTTTGAGACCCTGTCGTACCTGCCACCGCTCACGGTCGAGGACCTCTTGAAGCAGATCGAGTACCTGCTCCGGTCCAAATGGGTGCCTTGCCTCGAGTTT
** ***** ***** ******** ***** ** ** *** **** ** * ** ** ***** ** ********************** ** ** ** ******** *** * *****
RbcS4
RbcS1
RbcS2
RbcS3
AGCAAGGTCGGATTCGTCTACCGTGAGAATCACAGATCACCCGGATACTATGACGGTAGGTACTGGACCATGTGGAAACTGCCCATGTTTGGATGCACCGATGCCACCCAAGTGCTCAAG
AGCAAGGTCGGATTCGTCTACCGTGAGAACCACAGATCCCCCGGATACTACGATGGCAGGTACTGGACCATGTGGAAGCTGCCCATGTTCGGGTGCACTGACGCCACCCAGGTGCTCAAG
AGCAAGGTTGGATTCGTCTACCGTGAGAACCACAGGTCTCCCGGGTACTACGATGGCAGGTATTGGACCATGTGGAAGCTGCCCATGTTCGGCTGCACCGATGCCACCCAGGTGCTCAAG
AGTAAGGTCGGGTTCGTCTACCGTGAGAACCACAGGTCTCCTGGGTACTATGATGGCAGGTACTGGACCATGTGGAAGCTGCCTATGTTCGGATGCACTGATGCCACCCAGGTGCTCAAG
** ***** ** ***************** ***** ** ** ** ***** ** ** ***** ************** ***** ***** ** ***** ** ******** *********
RbcS4
RbcS1
RbcS2
RbcS3
GAGCTCGAGGAGGCAAAGAAGGCATACCCTGATGCCTTCGTTCGCATCATTGGCTTTGACAACGTTAGGCAGGTGCAGTTAATCAGCTTCATCGCGTATAAGCCACCGGGTTGCGAGGAG
GAGCTCGAGGAGGCCAAGAAGGCGTACCCTGATGCATTCGTCCGTATCATCGGCTTCGACAACGTCAGGCAGGTGCAGCTCATCAGCTTCATCGCCTACAAGCCCCCGGGCTGCGAGGAG
GAGCTCGAGGAGGCCAAGAAGGCCTACCCCGATGCCTTTGTCCGTATCATCGGCTTCGACAACGTCAGGCAGGTGCAGTTGATTAGCTTCATCGCCTACAAGCCCCCAGGTTGCGAGGAG
GAGCTCGAGGAGGCCAAGAAGGCATATCCAGATGCATTTGTTCGCATCATCGGCTTCGACAACGTCAGGCAGGTGCAGTTGATTAGCTTCATCGCCTACAAGCCCCCGGGTTGCGAGGAG
************** ******** ** ** ***** ** ** ** ***** ***** ******** ************ * ** *********** ** ***** ** ** *********
RbcS4
RbcS1
RbcS2
RbcS3
TCAGGCGGCAACTAAGCGCTTTCGTTCCTTCGTGCATGTTCTTTCTTTTTCTTTTTTTTTTGTGTGTCCGTGTTAAGCTGCACGTAATT-------GTTCTCTCGCGCTCCGACCTGCCG
TCTGGTGGCAACTAAGCCGTCATCGTCATATATAGCCTTGTTTAATTGTTCATCTCTGA----TTCG-----ATGATGTCTCCCACC--------TTGTTTCGTGTGTTCCCAGTTTG-TCTGGTGGCAACTAAGCTAAGATCAAGCATCGCGCTGGTGGATTGCTGCCTATAATAATAGTATGCAGCT--TTGTTTTGGGCTATG---------TTGATGATATATCAATATATAATA
TCCGGCGGAAACTAAGCTAGTTGGCAAGCCAGCAAGCTCACTCGTG-AGCTCTATACAGAGGAGACTCGA--TTGATCTATTCGGTTTTGGCAGCTTTGAACGTTCGAATCGGGCCACCT
** ** ** ********
*
*
*
*
RbcS4
RbcS1
RbcS2
RbcS3
TTGTTGCAAGAGTACTACTACAACTATCGGTCTATCGTTCGGTGACGGTGAGACAGG------------GCACGTGAATGCAAG--ATCTCCGGCTATACACACGTACTCATGTAATATG
TTCATCGTCTTTTGATTTTACCG--GCCGTGCTCTGCTTTTGTTTTTGTTTCACCTGATCTCTCTCTGACTTGATGTAAGAGTG--GTATCT---------------------------TGCTATATATTTTTATTTTACAG--TT----TGGTTATGTACCATCTCAATGGCCTCTGCTCTTAAC--ACATATGTAATAATCTCTTCCCTCCCTCTCCGGC----CGGTTTTATTGTA
TGCATTGGTTTCCCCTTTCCCAG--TT----TGTTGCTTCAGTTTTTTTTTTTCATCTTG----------TTTGTGTCAACATCAGCTGTGT--ATGTATGTA----CATATGTAT---*
*
*
* *
*
**
*
RbcS4
RbcS1
RbcS2
RbcS3
ATGCCTAGAGCATATCTGAATCCGTCGACAA---TGAAATTTTGGTTTTGCAAAATGCTGGTATTTGTTTATCATCCTGGCACGTGATATTTGCCTAGAGCATCTAAATCACTTTTACGA
--GCTACGACTATATGTTGTTGGGTGAGGCATATGTGAATGAAATATATGGAAGCTCCGGCTATATATATTTAT---------------------------------------------A-GAGTACTACAATTATCGTTGGG-TGAGGATATGTGAAAAC--------AAAGCTCCGGCTATATA------CACACA-----AGATATGTATCTAGAGCATATC----------------GTATCAACATAGGTTGAAGAG-CATGCATATGTGAATGC--------TAGCACCAGGCTAATCATATGCACATACACATCGTAATAAATATATTGTACAAATC-------------*
*
*
*
**
*
* **
RbcS4
RbcS1
RbcS2
RbcS3
AATGTGCGCGTCAACAAACTGATACGGCCCAAATGCCAGAAATTACCAGCATATATAGCCATATCAACTTTTGATTCGTATATATGAAGGTTGATTTAGTTAGAGAAATTCGGTTGTGAG
----------------------------------------------------------------------------------------------------------------------------------------------------------TGATAAATTCGT------------CG-------------------------------------------------------------GTCAAA--AACGATACTTATA---------TGTAATACTATACT------------CAGACC-AAACTTGATACCTCTGGTTTCC--------------------------------
RbcS4
RbcS1
RbcS2
RbcS3
AGAAGGAGGCTAGCAAAGATTCGGTTGATCAAGCTGTACCGCCAGGCCAGGACGTGCTGTGCGCGCGGCTGTGCCGCTTGACCGCAGAACCATACGGAATTGACGGACCAATTGTGCATA
----------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------------
Supplemental figure S1 cDNA alignment of the four RbcS genes using CLUSTALW (Larkin
et al. 2007). Both translation start and stop site were indicated by red bold letters
RbcS1
RbcS2
RbcS3
RbcS4
Supplemental figure S2 Expression profiles of the four rice RbcS gene in the developmental
stages and tissues. Plots were obtained from the spatio-temporal profiles of RiceXPro
(http://ricexpro.dna.affrc.go.jp/). X- axis shows various sources of tissues in rice and Y-axis
shows raw signal intensity of each gene in the tissues.