Supplementary File 1A PCR primers used in this study
Name
AP
Sequence (5’-3’)
GGCCACGCGTCGACTAGTACTTTTTTT
comments
Used for RT-PCR
TTTTTTTTTT
AUAP
GGCCACGCGTCGACTAGTAC
3’RACE for cloning
DP1
ATGKCAGASAMCAANMAGARTG
3’RACE for cloning
DP2
ATGKCWRRCAAYWSYCAGA
3’RACE for cloning
DP3
ATGGAYAAYARGCAAAACRYGAG
3’RACE for cloning
CbKIN1F
CCTTCCAAGCCGGTCAGA
3’RACE for 3’UTR cloning/qPCR
for gene expression
CbKIN1R
CCGATATGTTCTTTCCCGCT
qPCR for gene expression
CbKIN2F
CTTCCAAGCCGGTCAGG
3’RACE for 3’UTR cloning/qPCR
for gene expression
CbKIN2R
CGATATGTTCTTTCCCGCT
qPCR for gene expression
CbLEA1F
TGCCCAATCTGCTAAGGAA
3’RACE for 3’UTR cloning/qPCR
for gene expression
CbLEA1R
CAACGTGGTTCTCATTTGTTG
qPCR for gene expression
CbLEA2F
AATGCTGCCCAATCTGCC
3’RACE for 3’UTR cloning/qPCR
for gene expression
CbLEA2R
GTCTTCATTTGTTCATGCCAGTA
qPCR for gene expression
CbLEA3F
AAGGCAGGTGGTATGATGGA
qPCR for gene expression
CbLEA3R
AGGAAGTTGGAAGGGATTAGTG
qPCR for gene expression
CbLEA3F2
GACAACAGGCAAAACATGAGC
3’RACE for 3’UTR cloning
ACT7R
GAGCGATGGCTGGAATAGAA
qPCR for gene expression
ACT7F
GAGCGAGAAATTGTCCGTGA
qPCR for gene expression
Supplementary File 1B Alignment of amino acid sequences of KIN-related proteins from C.
bursa-pastoris and other plants
Supplementary File 1C Alignment of nucleotide sequences of the five KIN and KIN-homologous
LEA genes. Specific primer sets of CbKIN1 are marked by blue shaded background, CbKIN2 by
pink shaded background, CbLEA1 by red shaded background, CbLEA2 by yellow shaded
background, CbLEA3 by green shaded background. Initiation codons of the five genes are denoted
by single underline. Termination codons are denoted double underline
Supplementary File 1D Alignment of amino acid sequences of two KIN and three
KIN-homologous LEA proteins in C. bursa-pastoris using ClustalW2. Residues that are identical
in all sequences are shown with a black background. Partially conserved residues are shown with a
gray background
Supplementary File 1E The tree derived by neighbor-joining method with bootstrap analysis
(1000 replicates) from alignment of amino acid sequences of conserved region of KIN and
KIN-related proteins from C. bursa-pastoris and other plants. Only the corresponding amino acid
sequences were shown in Supplementary File 1B. Numbers above the major branches indicate
bootstrap values >50%. The abbreviations of species are as follows: Cb-Capsella bursa-pastoris,
At-Arabidopsis thaliana, Al-Arabidopsis lyrata, Bo-Brassica oleracea, Br-Brassica rapa,
Bn-Brassica napus, Rc-Ricinus communis, Gm-Glycine max, Cs-Camellia sinensis, Vv-Vitis
vinifera, Hs-Humulus scandens, Pt-Populus trichocarpa, Fs-Fagus sylvatica. The abbreviations of
protein are as follows: LEA (late embryogenesis abundant protein family protein), HYP
(hypothetical protein), PCP (pollen coat protein), AIP (ABA-inducible protein), CRP (cold
resistance protein), UNP (unknown protein). Two KIN and three KIN-homologous LEA proteins
of C. bursa-pastoris are shown in bold
Supplementary File 1F Variable 3’ UTRs of the five KIN-related genes were aligned using
CLUSTAL X (1.83). Termination codons of KIN-related genes are marked by green shaded
background
Four 3’UTR in CbKIN1
Four 3’UTR in CbKIN2
Four 3’UTR in CbLEA1
Four 3’UTR in CbLEA2
Two 3’UTR in CbLEA3
Supplementary File 1G qPCR showing CbLEA2 expression when plants were treated with 42 °C.
ACT7 was used as an internal control. The vertical column indicates the relative transcript level.
Data represent means ±standard deviation of two biological replicates