General characteristics of the G. guangxiense cp genome
7,428,469 high quality reads with a sequence length of 150 bp were used for the cp genome assembly, the raw reads have been deposited in NCBI SRA database under the accession No.PRJNA822055. The complete cp genome sequence of G. guangxiense was 157,785 bp in length (Fig. 1), consisting of a pair of inverted region (IR) regions (26,288 bp), a large single copy (LSC) region (86,702 bp) and a small single copy (SSC) region (18,507 bp). The universal GC content was 37.0%, however, the GC content was varied between different regions, specifically: 44.3% in the IR regions, 37.1% in the LSC region and 33.3% in the SSC region.
A total of 130 genes were found in the G. guangxiense cp genome, of which 113 are unique, including 79 protein-coding genes, 30 tRNA genes, and 4 rRNA genes (Fig. 1; Table 1). Among the 17 duplicated genes, there were 6 protein-coding genes (rpl2, rpl23, ycf2, ndhB, rps7, rps12), 7 tRNA genes(trnI-CAU, trnL-CAA, trnV-GAC, trnI-GAU, trnA-UGC, trnR-ACG, trnN-GUU) and 4 rRNA genes(rrn16, rrn23, rrn4.5, rrn5). The LSC region comprises 62 protein-coding and 22 tRNA genes, whereas the SSC region comprises 12 protein-coding genes and one tRNA gene. The protein-coding genes include nine genes encoding large ribosomal proteins (rpl2, rpl14, rpl16, rpl20, rpl22, rpl23 rpl32, rpl33, rpl36), 12 genes encoding small ribosomal proteins (rps2, rps3, rps4, rps7, rps8, rps11, rps12, rps14, rps15, rps16, rps18, rps19), five genes encoding photosystemⅠcomponents (psaA, psaB, psaC, psaI, psaJ), 15 genes related to photosystemⅡ(Table 1), and six genes (atpA, atpB, atpE, atpF, atpH, atpI) encoding ATP synthase and electron transport chain components (Table 1). 18 intron-containing genes were detected in the G. guangxiense cp genome, including nine protein-coding genes and six tRNA genes with one intron, three protein-coding genes (clpP, rps12, ycf3) had two intron (Table 2). We found that trnK-UUU had the largest intron (2,487 bp) and included the gene matK, the tRNA gene trnL-UAA had the smallest intron (540 bp).
Table 1 List of genes in the chloroplast genome of G. guangxiense
Category
|
Group of genes
|
Name of genes
|
Self-replication
|
Ribosomal proteins (Large subunit) (11)
|
1rpl2(×2), rpl14, 1rpl16, rpl20, rpl22, rpl23(×2), rpl32, rpl33, rpl36
|
|
Ribosomal proteins (small subunit )(14)
|
rps2, rps3, rps4, rps7(×2), rps8, rps11, 2rps12(×2), rps14, rps15,1 rps16, rps18, rps19
|
|
DNA dependent RNA polymerase(4)
|
rpoA, rpoB, 1rpoC1, rpoC2
|
|
Ribosomal RNAs (8)
|
rrn16(×2), rrn23(×2),rrn15(×2),rrn4.5(×2)
|
|
tRNA genes(37)
|
trnH-GUG,1trnk-UUU, trnQ-UUG, trnS-GCU,
trnG-UCC, trnR-UCU, trnC-GCA, trnD-GUC,
trnY-GUA, trnE-UUC, trnT-GGU, trnS-UGA,
1trnG-GCC, trnfM-CAU, trnS-GGA, trnT-UGU,
1trnL-UAA, trnF-GAA,1trnV-UAC, trnM-CAU,
trnW-CCA, trnP-UGG, trnI-CAU(×2), trnL-CAA(×2),
trnV-GAC(×2),1trnI-GAU(×2),1trnA-UGC(×2), trnR-ACG(×2), trnN-GUU(×2), trnL-UAG
|
Photosynthesis
|
PhotosystemI(5)
|
psaA, psaB, psaC, psaI, psaJ
|
|
PhotosystemII(15)
|
psbA, psbB, psbC, psbD, psbE, psbF, psbH, psbI, psbJ, psbK, psbL, psbM, psbN, psbT, psbZ
|
|
NadH oxidoreductase(12)
|
1ndhA,1ndhB(×2), ndhC, ndhD, ndhE, ndhF, ndhG, ndhH, ndhI, ndhJ, ndhK
|
|
Cytochrome b6/f complex(6)
|
petA, 1petB, 1petD, petG, petL, petN
|
|
ATP synthase(6)
|
atpA, atpB, atpE,1atpF, atpH, atpI
|
|
Rubisco large subunit(1)
|
rbcL
|
Other genes
|
Translation-related gene (1)
|
PinfA
|
|
Maturase(1)
|
matK
|
|
ATP-dependent protease subunit
p gene(1)
|
2clpP
|
|
Envelop membrane protein(1)
|
cemA
|
|
Acetyl-CoA carboxylase gene (1)
|
accD
|
|
C-type cytochrome synthesis gene(1)
|
ccsA
|
Unknown
|
Conserved open reading frames(5)
|
ycf1, ycf2(×2), 2ycf3, ycf4
|
Notes: 1: containing one intron; 2: containing two introns; P : pseudogenes; (×2): Gene with two copies
Repeat and SSR analysis
Repeat regions are essential in genome recombination and rearrangement (Nie et al. 2012). In this study, we searched for the types and distribution of repeated sequences and SSRs, the results showed that there were 62 repeats in the G. guangxiense cp genome, including 30 tandem repeats, 16 forward repeats, and 16 palindromic repeats (Table S1, S2). The tandem repeats were all between 11-32 bp long, the 16 forward repeats were 30-41 bp long, and the 16 palindromic repeats were 30-61 bp long (Table S2). The repeats distributed in intergenic spacer regions, protein-coding regions and above two regions were 54.84%(34 repeats), 25.81%(16 repeats) and 19.35%(12 repeats) respectively (Table S1, S2; Fig. 2A, B).
A total of 70 SSR loci were identified from G. guangxiense cp genome, including 44 mononucleotides (62.86%), 12 dinucleotides (17.14%), 2 trinucleotides (2.86%), 8 tetranucleotides (11.43%), 2 pentanucleotides (2.86%) and 2 hexanucleotides (2.86%). Among the 44 mononucleotides, 41 repeat units exhibited A or T types, 3 repeat units exhibited C and G type. The repeat number of mononucleotide motifs ranged from 10 to 17, but there is no repeats of 15 motifs. 53 of 70 SSR loci were found in the noncoding regions, the remaining 17 SSR loci were found in the gene coding regions (Table S3, S4; Fig. 2 C, D).
Table 2 Genes with introns in the chloroplast genome of G. Guangxiense
Gene
|
Location
|
Exon I (bp)
|
Intron I (bp)
|
Exon II (bp)
|
Intron II (bp)
|
Exon III (bp)
|
atpF
|
LSC
|
144
|
681
|
411
|
|
|
clpP
|
LSC
|
69
|
833
|
291
|
615
|
228
|
ndhA
|
SSC
|
552
|
1155
|
540
|
|
|
ndhB
|
IR
|
777
|
686
|
756
|
|
|
petB
|
LSC
|
6
|
780
|
642
|
|
|
petD
|
LSC
|
9
|
726
|
477
|
|
|
rpl16
|
LSC
|
9
|
1026
|
399
|
|
|
rpl2
|
IR
|
393
|
682
|
435
|
|
|
rpoC1
|
LSC
|
432
|
726
|
1602
|
|
|
rps12*
|
LSC
|
114
|
-
|
231
|
541
|
27
|
rps16
|
LSC
|
40
|
856
|
215
|
|
|
trnA-UGC
|
IR
|
38
|
802
|
35
|
|
|
trnG-UCC
|
LSC
|
20
|
659
|
51
|
|
|
trnI-GAU
|
IR
|
42
|
951
|
35
|
|
|
trnK-UUU
|
LSC
|
36
|
2487
|
36
|
|
|
trnL-UAA
|
LSC
|
38
|
540
|
48
|
|
|
trnV-UAC
|
LSC
|
39
|
601
|
37
|
|
|
ycf3
|
LSC
|
126
|
715
|
228
|
754
|
153
|
*rps12 gene is trans-spliced gene with the two duplicated 3’ end exons in IR regions and 5’ end exon in the LSC region.
Phylogenetic Analysis Based on Chloroplast Genome Sequence
To investigate the phylogenetic position of G. guangxiense, 73 shared protein-coding genes were used to conduct the MP and BI analyses, the selected models used in BI analyses were TVM+I+G. Both the MP and BI trees were well-supported and showed consistent phylogenetic patterns (Fig. 3; Table S5). It turned out that G. guangxiense has the closest phylogenetic relationship to G. caulopterum, and G. microspermum was the earliest diverging lineage in this group, all species of the Gynostemma clustered together to form a monophyletic clade.
Nucleotide composition between codon positions in the G. guangxiense cp genome
We screened 52 CDSs to be analyzed in G. guangxiense cp genome (Table 3), there were 20,844 codons (the termination codon were not included), and 61 kinds of codons encoding 20 animo acids. The four nucleotides distributed unevenly in the 52 CDSs, T (Thymine) had the highest mean value of 31.5%, A (adenine) had the second-highest value of 30.9%, followed by G (Guanine) and C (Cytosine), which was 20.3% and 17.4% respectively. To better estimate the nucleotide base composition in G. guangxiense, we caculated the CDS numbers with different GC content levels (Fig. 4A), the average value was 37.47% and the GC contents were in the range of 30.11–45.80%. The number of unigenes with GC contents of 40.0–41.0% was the largest group (the total number was 6), followed sequentially by the 35–36%, 39–40% and 43-44% intervals. The GC content in three codon positions (GC1, GC2, and GC3) were 45.68%, 37.66% and 29.07% respectively. This result suggested an obvious difference among three codon positions. GC3 was lower than GC1 and GC2, GC2 was similar to the average GC contents.
Table 3 GC content of different positions of each gene in the chloroplast genome of G. guangxiense
Genes
|
GC content (%)
|
ENC
|
CN
|
Genes
|
GC content (%)
|
ENC
|
CN
|
GC1
|
GC2
|
GC3
|
GCall
|
GC1
|
GC2
|
GC3
|
GCall
|
psbA
|
49.44
|
42.94
|
32.20
|
41.53
|
40.20
|
354
|
rpl20
|
35.29
|
41.18
|
25.74
|
34.07
|
44.64
|
136
|
matK
|
38.84
|
31.08
|
28.69
|
32.87
|
47.91
|
502
|
clpP
|
58.67
|
37.24
|
29.59
|
41.84
|
52.51
|
196
|
atpA
|
54.53
|
40.16
|
25.79
|
40.16
|
45.58
|
508
|
psbB
|
54.42
|
46.17
|
29.67
|
43.42
|
46.42
|
509
|
atpF
|
45.95
|
32.97
|
31.35
|
36.76
|
44.99
|
185
|
petB
|
49.07
|
41.67
|
30.56
|
40.43
|
45.51
|
216
|
atpl
|
49.19
|
36.69
|
25.00
|
36.96
|
45.55
|
250
|
petD
|
51.23
|
38.89
|
25.31
|
38.48
|
40.55
|
162
|
rps2
|
45.99
|
44.30
|
29.11
|
39.80
|
48.64
|
230
|
rpoA
|
44.88
|
31.33
|
26.81
|
34.34
|
51.16
|
332
|
rpoC2
|
46.12
|
36.64
|
27.80
|
36.85
|
50.10
|
1392
|
rps11
|
53.24
|
57.55
|
26.62
|
45.80
|
49.58
|
139
|
rpoC1
|
50.88
|
38.05
|
27.43
|
38.79
|
48.47
|
678
|
rps8
|
40.00
|
39.26
|
27.41
|
35.56
|
35.08
|
135
|
rpoB
|
49.30
|
38.38
|
27.45
|
38.38
|
48.60
|
1071
|
rpl14
|
52.85
|
38.21
|
21.14
|
37.40
|
43.81
|
123
|
psbD
|
52.26
|
43.22
|
31.92
|
42.47
|
44.21
|
354
|
rpl16
|
50.00
|
52.94
|
30.15
|
44.36
|
43.56
|
136
|
psbC
|
53.80
|
45.99
|
31.43
|
43.74
|
43.85
|
474
|
rps3
|
47.49
|
34.70
|
23.74
|
35.31
|
47.10
|
219
|
rps14
|
44.55
|
47.52
|
31.68
|
41.25
|
41.23
|
99
|
rpl22
|
37.13
|
34.13
|
32.34
|
34.53
|
45.86
|
167
|
psaB
|
48.57
|
42.72
|
29.39
|
40.23
|
46.56
|
737
|
rpl2
|
51.09
|
49.28
|
29.71
|
43.36
|
50.46
|
276
|
psaA
|
51.93
|
43.54
|
32.89
|
42.79
|
49.98
|
751
|
ycf2
|
41.45
|
34.18
|
36.72
|
37.45
|
52.79
|
2282
|
ycf3
|
47.34
|
38.46
|
33.14
|
39.64
|
61.00
|
169
|
ndhB
|
42.07
|
39.14
|
31.31
|
37.51
|
48.31
|
511
|
rps4
|
49.01
|
37.13
|
24.26
|
36.80
|
48.95
|
202
|
rps7
|
52.56
|
44.23
|
23.08
|
39.96
|
45.43
|
156
|
ndhJ
|
50.94
|
37.11
|
30.19
|
39.41
|
50.30
|
159
|
rps12
|
52.42
|
47.58
|
30.65
|
43.55
|
45.54
|
124
|
ndhK
|
42.29
|
45.37
|
26.87
|
38.18
|
49.79
|
227
|
ndhF
|
35.29
|
35.29
|
25.27
|
31.95
|
47.13
|
748
|
ndhC
|
48.76
|
31.40
|
25.62
|
35.26
|
46.07
|
121
|
ccsA
|
33.02
|
37.65
|
25.62
|
32.10
|
44.03
|
324
|
atpE
|
51.85
|
40.47
|
29.63
|
40.74
|
49.75
|
135
|
ndhE
|
39.60
|
33.66
|
26.73
|
33.33
|
54.50
|
101
|
atpB
|
55.91
|
41.48
|
28.06
|
41.82
|
45.34
|
489
|
ndhG
|
42.37
|
35.59
|
29.38
|
35.78
|
49.71
|
177
|
rbcL
|
58.19
|
43.70
|
29.62
|
43.84
|
48.19
|
476
|
ndhI
|
42.51
|
36.53
|
25.75
|
34.93
|
45.70
|
167
|
accD
|
39.23
|
37.80
|
28.66
|
35.23
|
46.45
|
492
|
ndhA
|
44.23
|
36.81
|
20.05
|
33.70
|
43.08
|
364
|
ycf4
|
42.08
|
41.53
|
37.16
|
40.26
|
56.95
|
183
|
ndhH
|
51.78
|
36.29
|
30.20
|
39.42
|
50.10
|
385
|
cemA
|
40.00
|
27.39
|
31.74
|
33.04
|
39.92
|
230
|
ycf1
|
36.31
|
28.50
|
25.54
|
30.11
|
47.02
|
1895
|
petA
|
52.80
|
36.96
|
32.30
|
40.68
|
52.43
|
322
|
total
|
45.68
|
37.66
|
29.07
|
37.47
|
49.24
|
|
rps18
|
35.29
|
40.20
|
26.47
|
33.99
|
38.77
|
102
|
|
|
|
|
|
|
|
Notes: GC1: GC value of first position of codon; GC2: GC value of second position of codon; GC3: GC value of third position of codon; GCall: GC value of each gene; ENC: efective number of codons.
The codon usage pattern of the G. guangxiense cp genome
The relative synonymous codon usage (RSCU) values of G. guangxiense unigenes are shown in Table 4. As reported, RSCU values are close to 1.0 when all synonymous codons are used equally without any bias, when RSCU≤1, it signifies no preference, 1<RSCU<1.2 indicates low preference, 1.2≤RSCU≤1.3 means moderate preference, and RSCU>1.3 represents high preference (Zuo et al. 2017). In our study, there were 30 codons with RSCU>1.0, of which, two codons exhibited low preference with 1<RSCU<1.2, five codons were median preference with 1.2≤RSCU≤1.3, and 23 codons showed strong prefernce with RSCU>1.3, the UUA encoding for arginine (Phe) was the highest RSCU value (1.97). In addition, it is worth mentioning that there were 29 codons ending with A/U among all 30 preferred codons.
Table 4. RSCU analysis of protein coding region in the chloroplast of G. guangxiense
Amino acid
|
codon
|
Number
|
RSCU
|
Amino acid
|
codon
|
Number
|
RSCU
|
Amino acid
|
codon
|
Number
|
RSCU
|
Phe
|
UUU
|
788
|
1.36
|
Thr
|
ACU
|
421
|
1.59
|
Glu
|
GAA
|
862
|
1.52
|
|
UUC
|
372
|
0.64
|
|
ACC
|
205
|
0.77
|
|
GAG
|
271
|
0.48
|
Leu
|
UUA
|
716
|
1.97
|
|
ACA
|
319
|
1.20
|
Cys
|
UGU
|
178
|
1.50
|
|
UUG
|
448
|
1.23
|
|
ACG
|
114
|
0.43
|
|
UGC
|
60
|
0.50
|
|
CUU
|
460
|
1.27
|
Ala
|
GCU
|
525
|
1.85
|
Arg
|
CGU
|
288
|
1.39
|
|
CUC
|
129
|
0.36
|
|
GCC
|
180
|
0.63
|
|
CGC
|
79
|
0.38
|
|
CUA
|
285
|
0.78
|
|
GCA
|
303
|
1.07
|
|
CGA
|
292
|
1.41
|
|
CUG
|
141
|
0.39
|
|
GCG
|
126
|
0.44
|
|
CGG
|
78
|
0.38
|
Ile
|
AUU
|
879
|
1.47
|
Tyr
|
UAU
|
626
|
1.57
|
Arg
|
AGA
|
367
|
1.78
|
|
AUC
|
341
|
0.57
|
|
UAC
|
171
|
0.43
|
|
AGG
|
135
|
0.65
|
|
AUA
|
571
|
0.96
|
His
|
CAU
|
388
|
1.54
|
Gly
|
GGU
|
500
|
1.39
|
Val
|
GUU
|
428
|
1.47
|
|
CAC
|
116
|
0.46
|
|
GGC
|
147
|
0.41
|
|
GUC
|
135
|
0.47
|
Gln
|
CAA
|
586
|
1.56
|
|
GGA
|
562
|
1.56
|
|
GUA
|
442
|
1.52
|
|
CAG
|
167
|
0.44
|
|
GGG
|
230
|
0.64
|
|
GUG
|
156
|
0.54
|
Pro
|
CCU
|
329
|
1.55
|
Asn
|
AAU
|
784
|
1.55
|
Ser
|
UCU
|
426
|
1.64
|
|
CCC
|
181
|
0.85
|
|
AAC
|
229
|
0.45
|
|
UCC
|
244
|
0.94
|
|
CCA
|
233
|
1.10
|
Lys
|
AAA
|
863
|
1.54
|
|
UCA
|
315
|
1.21
|
|
CCG
|
108
|
0.51
|
|
AAG
|
258
|
0.46
|
|
UCG
|
151
|
0.58
|
Asp
|
GAU
|
697
|
1.63
|
Met
|
AUG
|
473
|
1.00
|
|
AGU
|
326
|
1.25
|
|
GAC
|
159
|
0.37
|
TER
|
UAA
|
34
|
1.96
|
|
AGC
|
97
|
0.37
|
Trp
|
UGG
|
384
|
1.00
|
|
UAG
|
7
|
0.40
|
|
|
|
|
|
|
|
|
|
UGA
|
11
|
0.63
|
Neutrality plot analysis
Neutrality plots was used to estimate the degree of change in natural selection and mutation pressure by caculating the relationships between GC12 and GC3 (Sueoka 1988). In this study, the neutrality plot (Fig. 4B) reflected a weak correlation between GC12 and GC3, which suggested that codons usage were affected by mutation pressure to a very limited degree. Specifically, accoding to the slope of neutrality plot, mutation pressure only accounted for 17.46%, while natural selection accounted for 82.54%. These results suggested that natural selection played an important role in the pattern of G. guangxiense codon usage.
ENC plot analysis
In this study, ENC values of the different genes varied from 35.08-61.00, the average of which was 49.24, displaying different trends among the genes (Fig. 4C). Two cases were shown from the figure: one is some points on or very close to the curve, the other is some points far from the curve. The former indicated that the observed codon bias was due primarily to nucleotide composition bias at the third codon position (GC3S), the latter indicated that other factors like selection force also play an important role in codon usage choice among genes besides mutation force (Wright 1990). To obtain a more accurate estimation of the differences in ENC values, the results of (ENCexp-ENCobs)/ENCexp were calculated (Table S6). There were 21 genes distributed in the range of -0.05-0.05 and 31 genes distributed outside the -0.05-0.05 interval. This results reflected a great difference between ENCexp and ENCobs, most observed ENC values were lower than expected values, indicating that the codon use bias of G. guangxiense chloroplast genome was greatly affected by natural selection and less affected by mutation factors.
PR2-plot mapping analysis
According to PR2 plot mapping analysis result (Figure 4D), The distribution range of points in the plot is 0.35-0.61 in the Y-axis (A3 /(A3 + T3 ) and 0.28-0.67 in the X-axis (G3 /(G3 + C3). The genes were unevenly distributed in the four quadrants, Specifically, taking 0.5 as the boundary, there are more points on the right side of the X-axis, similarly, there were more points on the lower side of the Y-axis. The results showed that G and T were used more frequently than C and A in G. guangxiense, and the uneven distribution further suggested that the role of natural selection pressure is greater than that of mutation pressure in the codon uasge bias.
Identification of optimal codons
30 codons with the RSCU>1 were considered to be high frequency synonymous codons (Table 4). Meanwhile, 23 codons with △RSCU ≥ 0.08 were considered to be highly expressed codons (Table 5). Finally, 16 codons with a high frequency as well as high expression, including UUA、CUU、CGU、GCU、AUU、AAA、GUU、GUA、UCU、CCU、ACU、CAA、GAA、UGU、AGU、GGU were determined to be the optimal codons, of which, 11 ended with T and 5 ended with A. The results showed that the codons ending with C and G were lacked preference in the G. guangxiense cp genome.
Table 5 Putative optimal codons in the chloroplast genome of G. guangxiense
Amino acid
|
Codon
|
High expressed genes
|
Low expressed genes
|
△RSCU
|
Number
|
RSCU
|
Number
|
RSCU
|
Phe
|
UUU
|
31
|
1.07
|
108
|
1.08
|
-0.01
|
|
UUC
|
27
|
0.93
|
92
|
0.92
|
0.01
|
Leu
|
UUA***
|
35
|
1.89
|
55
|
1.08
|
0.81
|
|
UUG
|
24
|
1.30
|
75
|
1.47
|
-0.17
|
|
CUU**
|
27
|
1.46
|
76
|
1.49
|
-0.03
|
|
CUC
|
3
|
0.16
|
30
|
0.58
|
-0.42
|
|
CUA*
|
21
|
1.14
|
41
|
0.8
|
0.34
|
|
CUG
|
1
|
0.05
|
29
|
0.57
|
-0.52
|
Ile
|
AUU**
|
58
|
1.71
|
102
|
1.24
|
0.47
|
|
AUC
|
20
|
0.59
|
64
|
0.78
|
-0.19
|
|
AUA
|
24
|
0.71
|
80
|
0.98
|
-0.27
|
Met
|
AUG
|
27
|
1.00
|
62
|
1
|
0.00
|
Val
|
GUU*
|
22
|
1.54
|
41
|
1.38
|
0.16
|
|
GUC
|
4
|
0.28
|
17
|
0.57
|
-0.29
|
|
GUA***
|
27
|
1.89
|
37
|
1.24
|
0.65
|
|
GUG
|
4
|
0.28
|
24
|
0.81
|
-0.53
|
Ser
|
UCU***
|
29
|
2.60
|
70
|
1.51
|
1.09
|
|
UCC
|
10
|
0.90
|
55
|
1.19
|
-0.29
|
|
UCA
|
5
|
0.45
|
59
|
1.27
|
-0.82
|
|
UCG
|
4
|
0.36
|
37
|
0.8
|
-0.44
|
|
AGU*
|
11
|
0.99
|
40
|
0.86
|
0.13
|
|
AGC**
|
8
|
0.72
|
17
|
0.37
|
0.35
|
Pro
|
CCU***
|
19
|
1.69
|
32
|
1.17
|
0.52
|
|
CCC
|
9
|
0.80
|
27
|
0.99
|
-0.19
|
|
CCA
|
14
|
1.24
|
34
|
1.25
|
-0.01
|
|
CCG
|
3
|
0.27
|
16
|
0.59
|
-0.32
|
Thr
|
ACU**
|
20
|
1.60
|
37
|
1.22
|
0.38
|
|
ACC**
|
15
|
1.20
|
22
|
0.73
|
0.47
|
|
ACA
|
11
|
0.88
|
41
|
1.36
|
-0.48
|
|
ACG
|
4
|
0.32
|
21
|
0.69
|
-0.37
|
Ala
|
GCU***
|
35
|
2.55
|
43
|
1.83
|
0.72
|
|
GCC
|
3
|
0.22
|
18
|
0.77
|
-0.55
|
|
GCA
|
12
|
0.87
|
26
|
1.11
|
-0.24
|
|
GCG
|
5
|
0.36
|
7
|
0.3
|
0.06
|
Tyr
|
UAU
|
21
|
1.35
|
94
|
1.66
|
-0.31
|
|
UAC**
|
10
|
0.65
|
19
|
0.34
|
0.31
|
His
|
CAU
|
14
|
1.33
|
54
|
1.54
|
-0.21
|
|
CAC*
|
7
|
0.67
|
16
|
0.46
|
0.21
|
Gln
|
CAA*
|
19
|
1.65
|
82
|
1.4
|
0.25
|
|
CAG
|
4
|
0.35
|
35
|
0.6
|
-0.25
|
Asn
|
AAU
|
33
|
1.14
|
130
|
1.51
|
-0.37
|
|
AAC**
|
25
|
0.86
|
42
|
0.49
|
0.37
|
Lys
|
AAA**
|
31
|
1.59
|
104
|
1.27
|
0.32
|
|
AAG
|
8
|
0.41
|
60
|
0.73
|
-0.32
|
Asp
|
GAU
|
19
|
1.36
|
131
|
1.67
|
-0.31
|
|
GAC**
|
9
|
0.64
|
26
|
0.33
|
0.31
|
Glu
|
GAA**
|
41
|
1.58
|
111
|
1.28
|
0.30
|
|
GAG
|
11
|
0.42
|
62
|
0.72
|
-0.30
|
Cys
|
UGU***
|
6
|
2.00
|
23
|
1.35
|
0.65
|
|
UGC
|
0
|
0.00
|
11
|
0.65
|
-0.65
|
Trp
|
UGG
|
24
|
1.00
|
58
|
1
|
0.00
|
Arg
|
CGU***
|
19
|
2.07
|
25
|
0.77
|
1.30
|
|
CGC
|
4
|
0.44
|
13
|
0.4
|
0.04
|
|
CGA
|
14
|
1.53
|
49
|
1.52
|
0.01
|
|
CGG
|
1
|
0.11
|
17
|
0.53
|
-0.42
|
|
AGA
|
11
|
1.20
|
56
|
1.73
|
-0.53
|
|
AGG
|
6
|
0.65
|
34
|
1.05
|
-0.40
|
Gly
|
GGU***
|
41
|
2.38
|
38
|
1.09
|
1.29
|
|
GGC
|
6
|
0.35
|
16
|
0.46
|
-0.11
|
|
GGA
|
17
|
0.99
|
61
|
1.76
|
-0.77
|
|
GGG
|
5
|
0.29
|
24
|
0.69
|
-0.40
|
TER
|
UGA
|
1
|
0.60
|
1
|
0.6
|
0.00
|
TER
|
UAA
|
3
|
1.80
|
3
|
1.8
|
0.00
|
|
UAG
|
1
|
0.60
|
1
|
0.6
|
0.00
|
Note: “*”means ΔRSCU ≥ 0.08; “**”means ΔRSCU ≥ 0.3; “***”means ΔRSCU ≥ 0.5; “_” means the RSCU value is more than one; The codon in bold indicates that it is the optimal codon.