FOR ONLINE PUBLICATION ONLY
Supplementary Material 1
The specificity of degenerated primer pairs used for this study was demonstrated by DNA sequencing of the real-time quantitative RT-PCR products. The
degenerated primer pair used for the amplification is shown in column 1 (for more details see material section). The corresponding DNA sequencing results are
presented in column 2. The sequences were alignment against NCBI’s reference mRNA sequences (refseq_rna; blastn). Search was limited to Zea mays L.
(taxid:4577). Blast hits are presented in column 3 together with the respective Genbank accession numbers. Column 4 shows the resulting description of the
Blast hits. Parameters for statistical quality of the hit were presented in columns 5 - 7.
Significant alignments of sequences
Degenerated primer
pair used for real-time
quantitative RT-PCR
ZmMHA_fam
Sequencing summary of real-time
quantitative RT-PCR products
(using IUPAC base calls for wobbles)
Genbank accession
number
ACGCCTGGTCTGCTCCTGGTCACCGCGTTCC
TGCTCGCTCAACTTGTTGCCACCTTCCTC
NM_001112000.1
GCTGTCTACGCCAACTGGGGCTTCGCCAGGA
TCAAGGGTATCGGCTGGGGCTGGGCAGGT
GTGGTCTGGCTCTACAGCATCGTCSAT
TCKAGCCTGCGTGCAGCCGATGACTCCTTGC
TGTGCTTGTACGGCACGATGGACGCGGCG
ZmEGases
(NCBI blastn on Zea mays L. [taxid:4577]; database: refseq_rna)
CGGTGGTGCACCCGGCTGGGGTACTTGGGGC
CGTAGCCGACGAGGTAGCTGACGCCCCTG
GGGTTGGTGCCGAGCACGTAGTCCACCTGC
No other Blast hits
found
NM_001157964.1
No other Blast hits
found
Description
Zea mays plasma-membrane
H+ATPase2 (mha2), mRNA
>emb|X85805.1| Z.mays mRNA
for plasma membrane H+ ATPase
Zea mays endoglucanase 1
(LOC100285069), mRNA
>gb|EU970755.1| Zea mays
clone 350091 endoglucanase 1
precursor, mRNA, complete cds
Max score/
Total score
Query coverag.
(%)/ Max ident
(%)
E
value
207/ 207
95/ 93
1e53
263/ 263
93/ 99
2e70
Supplementary Material 2
a
b
ladder
1
Lector
2
3
1000 bp
900 bp
800 bp
700 bp
1
SR03
2
ladder
3
700 bp
600 bp
500 bp
500 bp
300 bp
200 bp
3
1
SR03
2
3
1000 bp
900 bp
800 bp
600 bp
400 bp
1
Lector
2
400 bp
300 bp
200 bp
100 bp
100 bp
Demonstration of the specificity of the degenerated primer pairs ZmMHA_fam and ZmEGases. After real-time quantitative RT-PCR measurement, PCR products
from both maize varieties were run on agarose gels. (a) By the use of the degenerated primer pair ZmMHA_fam for template amplification, only one single DNA
band was visualized. This band appeared at the molecular size that was predicted by in-silico analysis (expected size of PCR product of plasma membrane proton
pump (H+)–ATPase genes = 195 bp). (b) By the use of the degenerated primer pair ZmEGases, only one single DNA band was visualized. This band appeared at
the molecular size that was expected by in-silico analysis (predicted size of PCR product of endoglucanase genes = 200 bp). PCR conditions were as described in
Materials and Methods. Numbers 1-3, biological replicates of the real-time quantitative RT-PCR amplifications of Lector and SR03.Gels contain 1% agarose, Trisborate-EDTA, 0.4 µg of ethidium bromide per ml.