Biotechnology Bulletin ›› 2023, Vol. 39 ›› Issue (9): 105-116.doi: 10.13560/j.cnki.biotech.bull.1985.2023-0091
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LIN Hong-yan1,2,3(), GUO Xiao-rui1,2,3, LIU Di1,2,3, LI Hui1,2,3, LU Hai1,2,3()
Received:
2023-02-07
Online:
2023-09-26
Published:
2023-10-24
Contact:
LU Hai
E-mail:2638859438@qq.com;luhai1974@bjfu.edu.cn
LIN Hong-yan, GUO Xiao-rui, LIU Di, LI Hui, LU Hai. Molecular Mechanism of Transcriptional Factor AtbHLH68 in Regulating Cell Wall Development by Transcriptome Analysis[J]. Biotechnology Bulletin, 2023, 39(9): 105-116.
基因ID Unigene ID | 引物名称 Primer name | 引物序列 Primer sequence(5'-3') |
---|---|---|
AT3G18780 | Actin2-F | CGTATGAGCAAGGAGATCAC |
Actin2-R | CACATCTGTTGGAAGGTGCT | |
AT4G29100 | bHLH68-F | ACCACGGATGATGCCCACT |
bHLH68-R | CCATCATCAATCCACCCAAAA | |
AT1G56100 | PMEI-F | TGGTTGCCTCGTTGGATG |
PMEI-R | TGTGTAGGTTGACCATTGATGTAAA | |
AT2G47030 | VGD1-F | GCACAGTCCAGGTTGAGTCCGAAGG |
VGD1-R | GGTCTCCGTTCACTCTGATAGCCAC | |
AT4G34230 | CAD5-F | GTTGCCTCGTTGGATGTTGC |
CAD5-R | CGCCTTGTGTAGGTTGACCATTGAT | |
AT5G62350 | PMEI-F | ACTAAGCCGAGCCCAATCC |
PMEI-R | GTATCGTTCATCTCCTCGACGCAAT | |
AT3G56400 | WRKY70-F | GCGATAGTCGGAAAAGATTGG |
WRKY70-R | TGGGAATTTGGCATTAAGAA | |
AT1G30760 | BBE13-F | TCGGCGTAATACTTGCCTGGAAAAT |
BBE13-R | TATCAGCGACTTGTTGCCACTT | |
YXM-pER8-bHLH68-F | GGTAATGCCATGTAATATGCTCG | |
YXM-pER8-bHLH68-R | CATCATCAATCCACCCAAAA |
Table1 Primer sequence
基因ID Unigene ID | 引物名称 Primer name | 引物序列 Primer sequence(5'-3') |
---|---|---|
AT3G18780 | Actin2-F | CGTATGAGCAAGGAGATCAC |
Actin2-R | CACATCTGTTGGAAGGTGCT | |
AT4G29100 | bHLH68-F | ACCACGGATGATGCCCACT |
bHLH68-R | CCATCATCAATCCACCCAAAA | |
AT1G56100 | PMEI-F | TGGTTGCCTCGTTGGATG |
PMEI-R | TGTGTAGGTTGACCATTGATGTAAA | |
AT2G47030 | VGD1-F | GCACAGTCCAGGTTGAGTCCGAAGG |
VGD1-R | GGTCTCCGTTCACTCTGATAGCCAC | |
AT4G34230 | CAD5-F | GTTGCCTCGTTGGATGTTGC |
CAD5-R | CGCCTTGTGTAGGTTGACCATTGAT | |
AT5G62350 | PMEI-F | ACTAAGCCGAGCCCAATCC |
PMEI-R | GTATCGTTCATCTCCTCGACGCAAT | |
AT3G56400 | WRKY70-F | GCGATAGTCGGAAAAGATTGG |
WRKY70-R | TGGGAATTTGGCATTAAGAA | |
AT1G30760 | BBE13-F | TCGGCGTAATACTTGCCTGGAAAAT |
BBE13-R | TATCAGCGACTTGTTGCCACTT | |
YXM-pER8-bHLH68-F | GGTAATGCCATGTAATATGCTCG | |
YXM-pER8-bHLH68-R | CATCATCAATCCACCCAAAA |
Fig. 1 Screening of transgenic pER8-AtbHLH68 Arabidop-sis positive plant by PCR M:2 000 bp marker;1-19: pER8-AtbHLH68 transgenic positive plants. C1: Plasmid control. C2: Wild-type Arabidopsis. C3: Negative control
Fig. 2 Expression level of AtbHLH68 under estradiol treatments at different time points by RT-qPCR Control: DMSO treatment for 8 h. 2 h, 4 h, 6 h, 8 h: 10 μmol/L estradiol treatment of Arabidopsis stem for 2 h, 4 h, 6 h, 8 h. * P<0.05. ** P<0.01. *** P<0.001. The same below
样品名 Sample | 原始序列数 Raw reads/M | 高质量序列数 Clean reads/M | Q20/% | Q30/% | 比对到参考基因组序列数 No. of alignment reference genome sequences/% | 唯一参考序列数 No. of unique alignment sequences/% |
---|---|---|---|---|---|---|
4 h | 47.33 | 45.22 | 98.02 | 94.51 | 96.83 | 95.08 |
6 h | 45.57 | 43.95 | 97.83 | 93.99 | 97.12 | 95.35 |
8 h | 45.57 | 44.01 | 97.78 | 93.86 | 96.74 | 94.97 |
Control | 47.33 | 45 | 97.9 | 94.18 | 96.13 | 94.11 |
Table 2 Quality control data for transcriptome sequencing
样品名 Sample | 原始序列数 Raw reads/M | 高质量序列数 Clean reads/M | Q20/% | Q30/% | 比对到参考基因组序列数 No. of alignment reference genome sequences/% | 唯一参考序列数 No. of unique alignment sequences/% |
---|---|---|---|---|---|---|
4 h | 47.33 | 45.22 | 98.02 | 94.51 | 96.83 | 95.08 |
6 h | 45.57 | 43.95 | 97.83 | 93.99 | 97.12 | 95.35 |
8 h | 45.57 | 44.01 | 97.78 | 93.86 | 96.74 | 94.97 |
Control | 47.33 | 45 | 97.9 | 94.18 | 96.13 | 94.11 |
Fig. 3 4 h, 6 h and 8 h DEGs statistical number and Venn diagram A: 4 h volcano map of DEGs. B: 6 h volcano map of DEGs. C: 8 h volcano map of DEGs. D: 4 h, 6 h, 8 h Venn diagram of DGEs. Red dots: Up-regulated DEGs. Green dots: Down-regulated DEGs. Blue dots: Genes expressed without significant difference
基因表达Gene expression | 基因号Gene ID | Log2x | P值P Value | 描述Description |
---|---|---|---|---|
下调基因 | AT2G06850 | -1.06394221 | 1.85E-50 | Xyloglucan endotransglucosylase/hydrolase 4 |
Down-regulated gene | AT2G36870 | -1.266714752 | 3.34E-19 | Xyloglucan endotransglucosylase/hydrolase 32 |
AT4G25820 | -2.74819285 | 1.28E-06 | Xyloglucan endotransglucosylase/hydrolase 14 | |
AT5G57530 | -5.654911378 | 8.25E-28 | Xyloglucan endotransglucosylase/hydrolase 12 | |
AT5G57540 | -3.641717279 | 7.07E-17 | Xyloglucan endotransglucosylase/hydrolase 13 | |
AT1G71695 | -1.298988679 | 3.66E-22 | Peroxidase superfamily protein | |
AT1G30870 | -3.50779464 | 6.16E-08 | Peroxidase 7 | |
AT3G49110 | -2.420843121 | 6.80E-11 | Peroxidase 33 | |
AT3G50990 | -2.306246473 | 8.33E-06 | Peroxidase 36 | |
AT4G37980 | -1.756033681 | 3.73E-18 | Cinnamyl-alcohol dehydrogenase 7 | |
AT2G47030 | -2.29336778 | 2.45E-142 | Pectin methylesterase 4 | |
AT2G47040 | -2.786313785 | 0 | Vanguard1, VGD1 | |
AT3G10720 | -2.090197809 | 2.92E-13 | Pectin methylesterase 25 | |
AT3G62170 | -3.528498693 | 4.35E-207 | Pectin methylesterase 37 | |
AT4G15980 | -2.788495895 | 1.82E-05 | Pectin methylesterase 43 | |
AT5G07410 | -4.717215431 | 2.64E-240 | Pectin methylesterase 48 | |
AT3G05610 | -2.626956589 | 1.03E-120 | Pectin methylesterase 21 | |
AT2G26440 | -2.370550991 | 3.78E-132 | Pectin methylesterase 12 | |
AT3G06830 | -2.258402957 | 2.42E-15 | Plant invertase/pectin methylesterase inhibitor superfamily | |
AT5G07420 | -4.837943242 | 2.77E-08 | Pectin lyase-like superfamily protein | |
AT5G07430 | -2.606610505 | 7.12E-169 | Pectin lyase-like superfamily protein | |
AT3G17060 | -2.165538573 | 6.39E-22 | Pectin lyase-like superfamily protein | |
AT1G14420 | -2.298373367 | 3.91E-46 | Pectin lyase-like superfamily protein | |
AT2G02720 | -2.19730144 | 4.49E-24 | Pectin lyase-like superfamily protein | |
AT3G01270 | -2.71241092 | 0 | Pectin lyase-like superfamily protein | |
AT5G15110 | -2.336194664 | 2.35E-29 | Pectate lyase family protein | |
AT5G48900 | -6.312882955 | 1.10E-22 | Pectin lyase-like superfamily protein | |
AT5G55720 | -3.135159583 | 4.76E-11 | Pectin lyase-like superfamily protein | |
AT3G45860 | -1.803254412 | 3.34E-18 | Cysteine-rich RLK(receptor-like protein kinase)4 | |
AT4G23170 | -1.533408039 | 1.62E-109 | Cysteine-rich RLK(receptor-like protein kinase)9 | |
AT2G40750 | -1.152515323 | 8.63E-64 | WRKY DNA-binding protein 54 | |
AT2G46400 | -1.299919474 | 3.63E-61 | WRKY DNA-binding protein 46 | |
AT3G56400 | -1.40629482 | 0 | WRKY DNA-binding protein 70 | |
AT4G23810 | -1.452162691 | 1.12E-65 | WRKY DNA-binding protein 53 | |
AT5G24780 | -1.294335074 | 0 | Vegetative storage protein 1 | |
AT3G16470 | -1.212665758 | 0 | A JA-responsive gene that coordinates with GRP 7 | |
AT2G32690 | -2.181267688 | 6.01E-32 | Glycine-rich protein 23 | |
AT1G20510 | -1.035196623 | 4.70E-93 | OPC-8:CoA ligase involved in jasmonic acid biosynthesis | |
上调基因 | AT1G11545 | 2.054076673 | 1.34E-06 | Xyloglucan endotransglucosylase/hydrolase 8 |
Up-regulated gene | AT3G48580 | 3.262777493 | 3.34E-61 | Xyloglucan endotransglucosylase/hydrolase 11 |
AT4G14130 | 1.838579331 | 4.09E-18 | Xyloglucan endotransglucosylase/hydrolase 15 | |
AT4G37800 | 2.408582477 | 8.79E-21 | Xyloglucan endotransglucosylase/hydrolase 7 | |
AT1G30700 | 3.103198888 | 3.07E-254 | FAD-binding berberine family protein, AtBBE 7 | |
AT1G30720 | 3.04026387 | 2.09E-22 | FAD-binding berberine family protein, AtBBE 10 | |
AT1G30730 | 2.767165832 | 8.28E-09 | FAD-binding berberine family protein, AtBBE 11 | |
AT4G20800 | 3.940837928 | 8.47E-14 | FAD-binding berberine family protein, AtBBE 17 | |
AT5G44390 | 2.190683562 | 4.52E-05 | FAD-binding berberine family protein, AtBBE 25 | |
AT5G44400 | 2.779652634 | 0 | FAD-binding berberine family protein, AtBBE 26 | |
AT1G30760 | 1.250535773 | 3.26e-210 | Berberine bridge enzyme-like 13, AtBBE 13 | |
AT2G34790 | 2.036462643 | 1.06E-182 | Berberine bridge enzyme-like 15, AtBBE 15 | |
AT4G36430 | 2.677849798 | 3.42E-36 | Peroxidase superfamily protein | |
AT3G21770 | 1.766366511 | 5.33E-07 | Peroxidase superfamily protein | |
AT3G28200 | 1.80094768 | 1.06E-34 | Peroxidase superfamily protein | |
AT1G05260 | 1.600816452 | 2.62E-31 | Peroxidase 3 | |
AT5G05340 | 2.40141963 | 1.48E-80 | Peroxidase 52 | |
AT5G60020 | 1.132349101 | 5.96E-135 | Laccase 17 | |
AT4G34230 | 1.168213629 | 0 | Cinnamyl alcohol dehydrogenase 5 | |
AT4G37990 | 1.401205226 | 3.90E-06 | Cinnamyl-alcohol dehydrogenase 8 | |
AT1G11590 | 2.950611973 | 1.48E-19 | Pectin methylesterase 19 | |
AT5G09760 | 1.08363098 | 2.08E-22 | Pectin methylesterase 15 | |
AT1G14890 | 1.984238146 | 0 | Plant invertase/pectin methylesterase inhibitor superfamily protein | |
AT5G62350 | 1.901654358 | 0 | Plant invertase/pectin methylesterase inhibitor superfamily protein | |
AT1G62770 | 1.620506618 | 3.20E-05 | Pectin methyleseterase inhibitor 19 | |
AT1G56100 | 1.25 | 0 | Plant invertase/pectin methylesterase inhibitor 14 | |
AT3G47380 | 1.787395021 | 2.75E-23 | Plant invertase/pectin methylesterase inhibitor 11 | |
AT3G27400 | 2.985665084 | 5.10E-106 | Pectin lyase-like superfamily protein | |
AT3G53190 | 1.145912382 | 1.86E-14 | Pectin lyase-like superfamily protein | |
AT4G13710 | 1.186011986 | 2.50E-09 | Pectin lyase-like superfamily protein | |
AT4G24780 | 1.1622914 | 1.26E-84 | Pectate lyase 19 | |
AT5G04310 | 1.318331844 | 6.35E-11 | Pectate lyase like 12 |
Table 3 Functional analysis of DEGs
基因表达Gene expression | 基因号Gene ID | Log2x | P值P Value | 描述Description |
---|---|---|---|---|
下调基因 | AT2G06850 | -1.06394221 | 1.85E-50 | Xyloglucan endotransglucosylase/hydrolase 4 |
Down-regulated gene | AT2G36870 | -1.266714752 | 3.34E-19 | Xyloglucan endotransglucosylase/hydrolase 32 |
AT4G25820 | -2.74819285 | 1.28E-06 | Xyloglucan endotransglucosylase/hydrolase 14 | |
AT5G57530 | -5.654911378 | 8.25E-28 | Xyloglucan endotransglucosylase/hydrolase 12 | |
AT5G57540 | -3.641717279 | 7.07E-17 | Xyloglucan endotransglucosylase/hydrolase 13 | |
AT1G71695 | -1.298988679 | 3.66E-22 | Peroxidase superfamily protein | |
AT1G30870 | -3.50779464 | 6.16E-08 | Peroxidase 7 | |
AT3G49110 | -2.420843121 | 6.80E-11 | Peroxidase 33 | |
AT3G50990 | -2.306246473 | 8.33E-06 | Peroxidase 36 | |
AT4G37980 | -1.756033681 | 3.73E-18 | Cinnamyl-alcohol dehydrogenase 7 | |
AT2G47030 | -2.29336778 | 2.45E-142 | Pectin methylesterase 4 | |
AT2G47040 | -2.786313785 | 0 | Vanguard1, VGD1 | |
AT3G10720 | -2.090197809 | 2.92E-13 | Pectin methylesterase 25 | |
AT3G62170 | -3.528498693 | 4.35E-207 | Pectin methylesterase 37 | |
AT4G15980 | -2.788495895 | 1.82E-05 | Pectin methylesterase 43 | |
AT5G07410 | -4.717215431 | 2.64E-240 | Pectin methylesterase 48 | |
AT3G05610 | -2.626956589 | 1.03E-120 | Pectin methylesterase 21 | |
AT2G26440 | -2.370550991 | 3.78E-132 | Pectin methylesterase 12 | |
AT3G06830 | -2.258402957 | 2.42E-15 | Plant invertase/pectin methylesterase inhibitor superfamily | |
AT5G07420 | -4.837943242 | 2.77E-08 | Pectin lyase-like superfamily protein | |
AT5G07430 | -2.606610505 | 7.12E-169 | Pectin lyase-like superfamily protein | |
AT3G17060 | -2.165538573 | 6.39E-22 | Pectin lyase-like superfamily protein | |
AT1G14420 | -2.298373367 | 3.91E-46 | Pectin lyase-like superfamily protein | |
AT2G02720 | -2.19730144 | 4.49E-24 | Pectin lyase-like superfamily protein | |
AT3G01270 | -2.71241092 | 0 | Pectin lyase-like superfamily protein | |
AT5G15110 | -2.336194664 | 2.35E-29 | Pectate lyase family protein | |
AT5G48900 | -6.312882955 | 1.10E-22 | Pectin lyase-like superfamily protein | |
AT5G55720 | -3.135159583 | 4.76E-11 | Pectin lyase-like superfamily protein | |
AT3G45860 | -1.803254412 | 3.34E-18 | Cysteine-rich RLK(receptor-like protein kinase)4 | |
AT4G23170 | -1.533408039 | 1.62E-109 | Cysteine-rich RLK(receptor-like protein kinase)9 | |
AT2G40750 | -1.152515323 | 8.63E-64 | WRKY DNA-binding protein 54 | |
AT2G46400 | -1.299919474 | 3.63E-61 | WRKY DNA-binding protein 46 | |
AT3G56400 | -1.40629482 | 0 | WRKY DNA-binding protein 70 | |
AT4G23810 | -1.452162691 | 1.12E-65 | WRKY DNA-binding protein 53 | |
AT5G24780 | -1.294335074 | 0 | Vegetative storage protein 1 | |
AT3G16470 | -1.212665758 | 0 | A JA-responsive gene that coordinates with GRP 7 | |
AT2G32690 | -2.181267688 | 6.01E-32 | Glycine-rich protein 23 | |
AT1G20510 | -1.035196623 | 4.70E-93 | OPC-8:CoA ligase involved in jasmonic acid biosynthesis | |
上调基因 | AT1G11545 | 2.054076673 | 1.34E-06 | Xyloglucan endotransglucosylase/hydrolase 8 |
Up-regulated gene | AT3G48580 | 3.262777493 | 3.34E-61 | Xyloglucan endotransglucosylase/hydrolase 11 |
AT4G14130 | 1.838579331 | 4.09E-18 | Xyloglucan endotransglucosylase/hydrolase 15 | |
AT4G37800 | 2.408582477 | 8.79E-21 | Xyloglucan endotransglucosylase/hydrolase 7 | |
AT1G30700 | 3.103198888 | 3.07E-254 | FAD-binding berberine family protein, AtBBE 7 | |
AT1G30720 | 3.04026387 | 2.09E-22 | FAD-binding berberine family protein, AtBBE 10 | |
AT1G30730 | 2.767165832 | 8.28E-09 | FAD-binding berberine family protein, AtBBE 11 | |
AT4G20800 | 3.940837928 | 8.47E-14 | FAD-binding berberine family protein, AtBBE 17 | |
AT5G44390 | 2.190683562 | 4.52E-05 | FAD-binding berberine family protein, AtBBE 25 | |
AT5G44400 | 2.779652634 | 0 | FAD-binding berberine family protein, AtBBE 26 | |
AT1G30760 | 1.250535773 | 3.26e-210 | Berberine bridge enzyme-like 13, AtBBE 13 | |
AT2G34790 | 2.036462643 | 1.06E-182 | Berberine bridge enzyme-like 15, AtBBE 15 | |
AT4G36430 | 2.677849798 | 3.42E-36 | Peroxidase superfamily protein | |
AT3G21770 | 1.766366511 | 5.33E-07 | Peroxidase superfamily protein | |
AT3G28200 | 1.80094768 | 1.06E-34 | Peroxidase superfamily protein | |
AT1G05260 | 1.600816452 | 2.62E-31 | Peroxidase 3 | |
AT5G05340 | 2.40141963 | 1.48E-80 | Peroxidase 52 | |
AT5G60020 | 1.132349101 | 5.96E-135 | Laccase 17 | |
AT4G34230 | 1.168213629 | 0 | Cinnamyl alcohol dehydrogenase 5 | |
AT4G37990 | 1.401205226 | 3.90E-06 | Cinnamyl-alcohol dehydrogenase 8 | |
AT1G11590 | 2.950611973 | 1.48E-19 | Pectin methylesterase 19 | |
AT5G09760 | 1.08363098 | 2.08E-22 | Pectin methylesterase 15 | |
AT1G14890 | 1.984238146 | 0 | Plant invertase/pectin methylesterase inhibitor superfamily protein | |
AT5G62350 | 1.901654358 | 0 | Plant invertase/pectin methylesterase inhibitor superfamily protein | |
AT1G62770 | 1.620506618 | 3.20E-05 | Pectin methyleseterase inhibitor 19 | |
AT1G56100 | 1.25 | 0 | Plant invertase/pectin methylesterase inhibitor 14 | |
AT3G47380 | 1.787395021 | 2.75E-23 | Plant invertase/pectin methylesterase inhibitor 11 | |
AT3G27400 | 2.985665084 | 5.10E-106 | Pectin lyase-like superfamily protein | |
AT3G53190 | 1.145912382 | 1.86E-14 | Pectin lyase-like superfamily protein | |
AT4G13710 | 1.186011986 | 2.50E-09 | Pectin lyase-like superfamily protein | |
AT4G24780 | 1.1622914 | 1.26E-84 | Pectate lyase 19 | |
AT5G04310 | 1.318331844 | 6.35E-11 | Pectate lyase like 12 |
Fig. 6 Biosynthetic pathway map A: Pentose and glucuronate interconversions pathway map and DEG heat map, values are log2(FPKM+1). PME: Pectin methylesterase. PMEI: Pectin methylesterase inhibitor. PG: Polygalacturonase. PL: Pectate lyase. B: Phenylpropane biosynthesis pathway pathway map and DEG heat map, values are log2(FPKM+1). CAD: Cinnamyl alcohol dehydrogenase. PER: Peroxidase. LAC: Laccase
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