雄蕊發育相關基因之差異表現 - 苦瓜花性之轉錄體分析

將雄蕊發育相關基因整理如表 17。XERICO 之苦瓜同源基因 McXERICO.1 (BG17271.1、BG17271.2、BG17271.3) 及 McXERICO.2 (BG15458.1、BG15458.2) 於外施激勃素後表現量上升 (圖 16、表 17)。LOX1 為茉莉酸生合成相關基因，其 苦瓜之同源基因 McLOX1.2 (BG19644.1)、McLOX1.3 (BG8248.1) 及 McLOX1.4 (BG11124.1) 表現量同樣受激勃素處理而上升 (圖 16、表 17)。茉莉酸訊息傳遞相 關基因 McJAZ8 (BG28497.1) 與 McMYB24.1 (BG46078.1) 皆於激勃素處理後表現 量下降。編碼 ARF6 之基因 (BG42471.1、BG42465.1、BG19620.1、BG19619.1、

BG19618.1) 及 ARF8 之基因 (BG42468.1、BG42473.1)，於激勃素處理後表現量下降；編碼核轉錄因子 (nuclear transcription factors) EIN3/EILs 之基因於外施激勃素 後表現量則增加 (圖 16)。McAP3、McPI、McAG 為 MADS-box 之花器發育同源異 型基因，其中 McAP3 (BG1411.1) 及 McAG (BG30120.1) 於外施激勃素後表現量下 降，而 McPI (BG11587.1) 則於處理後表現量增加 (圖 16、表 17)。MYB33、MYB65 及 MYB101 為 GAMYB 同源基因，其中 MYB65 及 MYB101 於苦瓜無同源序列，而 McMYB33 (BG34956.1、BG34956.3、BG34956.4、BG34956.5) 則於激勃素處理後表 現量無顯著差異 (圖 16、表 17)。DAD1、MYB65、MYB101、MYB108、RGL3、JAZ1、

JAZ11 於苦瓜沒有比對到完整或部分開放解讀框架註解序列。

圖 1. 轉錄體次世代定序分析流程

Fig. 1. The NGS analysis workflow for the whole-transcriptome analysis pipeline.

Control and GA treatment samples from Momordica charantia ‘Moon Shine’ were selected for whole-transcriptome analysis using Illumina. The 106,055 de novo assembled M. charantia contigs sequences were compared with the TAIR and EMBL coding sequence (CDS) databases through BLAST for open reading frame (ORF) annotation. FPKM, Fragments per kilobase of transcript per million mapped reads. GO, Gene ontology.

圖 2. 開放解讀框架註解情形

Fig. 2. The characterization of open reading frame (ORF) annotations. (A) Pie chart showing the percentages of complete genes, partial genes and undefined contigs. (B) Fold change distribution between expression levels of control and GA treated bitter gourd seedlings for complete and partial ORF contigs.

圖 3. 差異性表現基因之生物過程功能分類

Fig. 3. Functional categorization of differentially expressed contigs after GA3 treatment based on the biological process of Gene Ontology.

圖 4. 差異性表現基因之分子功能分類

Fig. 4. Functional categorization of differentially expressed contigs after GA3 treatment based on the molecular function of Gene Ontology.

圖 5. 差異性表現基因之細胞組成分類

Fig. 5. Functional categorization of differentially expressed contigs after GA3 treatment based on the cellular component of Gene Ontology.

圖 6. 激勃素生合成途徑基因表現差異

Fig. 6. Representative gene expression profiles of the gibberellin biosynthesis pathway in Momordica charantia. The two color boxes represent gene expression level in control (C) and GA treated (T) bitter gourd. Log2 fold change was correlated to control.

The color scale is shown at the bottom. Red and green colors indicate higher and lower expression, respectively. Gray color indicates that no transcript was detected in GA3 -treated samples.The gibberellin biosynthesis pathway was referenced to the pathway in Arabidopsis (Fleet et al., 2003). Gray words mean no contigs align to the reference genes. CPS, CPP synthetase; KS, ent-kaurene synthase. KO1, ent-kaurene oxidase 1;

KAO, ent-kaurenoic acid oxydase; GA13ox, gibberellin 13-oxidase; GA20ox, gibberellin 20-oxidase; GA3ox, gibberellin 3-oxidase; GA2ox, gibberellin 2-oxidase.

圖 7. 激勃素訊息傳遞途徑基因表現差異

Fig. 7. Representative gene expression profiles of the gibberellin signal transduction pathway in Momordica charantia. The two color boxes represent gene expression level in control (C) and GA treated (T) bitter gourd. Log2 fold change was correlated to control. The color scale is shown at the bottom. Red and green colors indicate higher and lower expression, respectively. The gibberellin signal transduction pathway was referenced to the pathway in Arabidopsis (Wang and Deng, 2011). Gray words mean no contigs align to the reference genes. GID1, GA insensitive dwarf 1; GAI, GA insensitive; RGA, repressor or gai-3. RGL, RGA-like.

圖 8. 乙烯生合成途徑基因表現差異

Fig. 8. Representative gene expression profiles of the ethylene biosynthesis pathway in Momordica charantia. The two color boxes represent gene expression level in control (C) and GA treated (T) bitter gourd. Log2 fold change was correlated to control. The color scale is shown at the bottom. Red and green colors indicate higher and lower expression, respectively. Gray color indicates that no transcript was detected in GA3 -treated samples. The ethylene biosynthesis pathway was referenced to the pathway in Arabidopsis (Sauter et al., 2013). SAM, S-adenosylmethionine synthetase; ACS, ACC stnthase; ACO, ACC oxidase.

1 100 FB801118 (1) ATGGCAATGTTATCTAGCAAAGCTAGCCATGATTCTCATGGCCAGGACTCCTCCTACTTCTTTGGGTGGCAAGAGTACGAGAAGAACCCCTACCACCCAA McACS8.2 (1) --- McACS11 (1) --- 101 200 FB801118 (101) CTCGTAACCCCACTGGAATTATTCAAATGGGTCTCGCTGAAAACGAGCTGTCTTTCGACCTGGTGGAGGAGTGGCTGGATAGAAATCCAGATGCCTTGGG McACS8.2 (1) --- McACS11 (1) --- 201 300 FB801118 (201) ATTGAGAAGAAATGGGAAGTCAGTGTTCAGAGAGCTGGCGCTTTTCCAAGACTACCATGGCTTGCCCGCTTTTAAAAAAGAGCTGGTGGAGTTGATGGCG McACS8.2 (1) --- McACS11 (1) --- 301 400 FB801118 (301) GAAATACGAGGAAACAAAGTGAAATTCGATGTCAACAACCTCGTCCTCACCGCTGGTGCAACTTCTGCCAATGAGATCCTTATGTTTTGTCTCGCTCAAT McACS8.2 (1) ---TGAGATCCTTATGTTTTGTCTCGCTCAAT McACS11 (1) --- 401 500 FB801118 (401) CTGGAGAAGCCTTTCTCGTTCCCACCCCATACTATCCAGGGTTTGACAGGGACGTAAAATGGCGAACAGGGGCACAAATAATTCCAATCCAGTGTCCGAG McACS8.2 (30) CTGGAGAAGCCTTTCTCGTTCCCACCCCATACTATCCAGGGTTTGACAGGGACGTAAAATGGCGAACAGGGGCACAAATAATTCCAATCCAGTGTCCGAG McACS11 (1) --- 501 600 FB801118 (501) TTCAAACGGATTCCGAATCAGCGGTTGGGCCATGGAGGAGGCGTGGGAACGAGCCGAGGAGCTGAAACTGAGAGTGAAAGGGGTTCTAATAACGAACCCC McACS8.2 (130) TTCAAACGGATTCCGAATCAGCGGTTGGGCCATGGAGGAGGCGTGGGAACGAGCCGAGGAGCTGAAACTGAGAGTGAAAGGGGTTCTAATAACGAACCCC McACS11 (1) --- 601 700 FB801118 (601) TCCAACCCATTGGGCACCACAATGGGCCGCGACGAGCTCAATTTACTTGTGGACTTCGCCGCCGCCAAAGGCATCCACATCGTCAGCGACGAGATTTACT McACS8.2 (230) TCCAACCCATTGGGCACCACAATGGGCCGCGACGAGCTCAATTTACTTGTGGACTTCGCCGCCGCCAAAGGCATCCACATCGTCAGCGACGAGATTTACT McACS11 (1) --- 701 800 FB801118 (701) CCGCCACCGTCTTCGACTCCCCCTCTTTCATAAGCATCACCGAAGCCCTTATCGATCGGAACCTCCAAAATTCCCCACTTTGGAACCGAATTCATGTCGT McACS8.2 (330) CCGCCACCGTCTTCGACTCCCCCTCTTTCATAAGCATCACCGAAGCCCTTATCGATCGGAACCTCCAAAATTCCCCACTTTGGAACCGAATTCATGTCGT McACS11 (1) --- 801 900 FB801118 (801) CTATAGCCTCTCCAAGGACCTCGGAGTCCCCGGGTTCAGAGTGGGCATGATTTATTCCAACGACCGACATGTCGTCGCAGCGGCCACCAAAATGTCCAGC McACS8.2 (430) CTATAGCCTCTCCAAGGACCTCGGAGTCCCCGGGTTCAGAGTGGGCATGATTTATTCCAACGACCGACATGTCGTCGCAGCGGCCACCAAAATGTCCAGC McACS11 (1) --- 901 1000 FB801118 (901) TTCGGCCTCATCTCGTCGCAGACGCAGTACCTGCTGTCGCGGATGCTGTCGGACCGGGACTTCCGGGGGAATTATATGGACGAGACCAAGAGGCGGATCC McACS8.2 (530) TTCGGCCTCATCTCGTCGCAGACGCAGTACCTGCTG--- McACS11 (1) ---TCGCAGACGCAGTACCTGCTGTCGCGGATGCTGTCGGACCGGGACTTCCGGGGGAATTATATGGACGAGACCAAGAGGCGGATCC 1001 1100 FB801118 (1001) GGAAGAGAAAGGGGATGCTTGTTTCGGGGCTCCGGAACGCCGGAATCGGGTGCTTGGACAGCAATTCGGGTCTTTTCTGCTGGGTGGACATGAGGCATCT McACS8.2 (566) --- McACS11 (86) GGAAGAGAAAGGGGATGCTTGTTTCGGGGCTCCGGAACGCCGGAATCGGGTGCTTGGACAGCAATTCGGGTCTTTTCTGCTGGGTGGACATGAGGCATCT 1101 1200 FB801118 (1101) CTTAAAAAACGCCACTTTCGAGGACGAAATGGAGCTGTGGAGGACCATCTTGTGCCAGGTTGGGCTCAATATCTCCCCCGGTTCGGCTTGCCACTGCTCC McACS8.2 (566) --- McACS11 (186) CTTAAAAAACGCCACTTTCGAGGACGAAATGGAGCTGTGGAGGACCATCTTGTGCCAGGTTGGGCTCAATATCTCCCCCGGTTCGGCTTGCCACTGCTCC 1201 1300 FB801118 (1201) GAACCGGGTTGGTTCAGAATGTGCTTTGCCAATATGTCGGAACATACTCTCATGCTCGCGATAAGTCGTCTCAAGACATTCGTCGAATCCTCCTCTTCCG McACS8.2 (566) --- McACS11 (286) GAACCGGGTTGGTTCAGAATGTGCTTTGCCAATATGTCGGAACATACTCTCATGCTCGCGATAAGTCGTCTCAAGACATTCGTCGAATCCTCCTCTTCCG 1301 1400 FB801118 (1301) GTGGCGACGCTGATGCCAACGAGGAGTTCTGTCGAAGTAATCAGAAAGTTGTTGGTAGATCATACGGTAGGAAAAGGCTGTTAAGACGACTGCTTGAATC McACS8.2 (566) --- McACS11 (386) GTGGCGACGCTGATGCCAACGAGGAGTTCTGTCGAAGTAATCAGAAAGTTGTTGGTAGATCATACGGTAGGAAAAGGCTGTTAAGACGACTGCTTGAATC 1401 1442

FB801118 (1401) GGGGGTTTGTTCTTTGATGTATACGTGTTTTCCATGCTGA-- McACS8.2 (566) --- McACS11 (486) GGGGGTTTGTTCTTTGATGTATACGTGTTTTCCATGCTGAGC

圖9. 苦瓜McACS8.2及McACS11與FB801118之核苷酸序列比對

Fig. 9. Alignment of Momordica charantia McACS8.2, McACS11 and FB801118 nucleotide sequence. The black and gray boxes represent identical and conserved nucleotides, respectively.

1 100 BG42284.12 (1) ---ATGGTACTATTTGACATTC---- BG42288.1 (1) ATGGCAATCCCCATTATTGACTTCTCAAAGCTTAATGGAGAAGAAAGAGCCAATACATTGGCTCAGATAGCCAATGGCTGTGAAGAATGGGGATTCTTTC BG42289.1 (1) --- 101 200 BG42284.12 (20) AGCTGGTGAACCATGGGATTCCTGAGGAACTCTTGGAGAGGGTGAAGAGGGTGTGTTCAGAGTGCTACAAGCTGGAAAGAGAAGAGAGTTTCAAGAGTTC BG42288.1 (101) AGCTGGTGAACCATGGGATTCCTGAGGAACTCTTGGAGAGGGTGAAGAGGGTGTGTTCAGAGTGCTACAAGCTGGAAAGAGAAGAGAGTTTCAAGAGTTC BG42289.1 (1) --- 201 300 BG42284.12 (120) AAAACCTGTGAAGCTTCTGAATGATTTGCTGGAAAACAAGAGCGGTGAGAAGCTGGAGAATCTTGATTGGGAAGATGTCTTCCTTCTCCATGACGAAAAC BG42288.1 (201) AAAACCTGTGAAGCTTCTGAATGATTTGCTGGAAAACAAGAGCGGTGAGAAGCTGGAGAATCTTGATTGGGAAGATGTCTTCCTTCTCCATGACGAAAAC BG42289.1 (1) --- 301 400 BG42284.12 (220) GACTGGCCCTCCAACATTCCTG--- BG42288.1 (301) GACTGGCCCTCCAACATTCCTGGATTCAAGGAAACCATGGCAGAGTACAGATCGGAGCTGAAGAAGCTAGCAAAAAAGGTGATGGCAGTGATGGATGAGA BG42289.1 (1) --- 401 500 BG42284.12 (242) ---AGGGATACATAAACGCTGCCTTCAATGGTGGCGAAGGGCTGGAGAAAGCCTTCTTTGGCACCAAAGTCAGCCACTACCCACCATG BG42288.1 (401) ACTTAGGTTTACCTGAGGGATACATAAACGCTGCCTTCAATGGTGGCGAAGGGCTGGAGAAAGCCTTCTTTGGCACCAAAGTCAGCCACTACCCACCATG BG42289.1 (1) --- 501 600 BG42284.12 (327) CCCACACCCAGAGCTGGTGAACGGCCTCCGTGCACACACTGATGCTGGCGGTGTCATCTTGCTCTTCCAAGACGACCAGGTGGGAGGCCTTCAGATCCTG BG42288.1 (501) CCCACACCCAGAGCTGGTGAACGGCCTCCGTGCACACACTGATGCTGGCGGTGTCATCTTGCTCTTCCAAGACGACCAGGTGGGAGGC--- BG42289.1 (1) --- 601 700 BG42284.12 (427) AAAGATGGGCAGTGGATAGATGTGCAGCCATTACCCAACTCCATAGTCATCAACACAGGTGACCAGATTGAGGTCCTGAGCAATGGAAGATACAAAAGTG BG42288.1 (589) --- BG42289.1 (1) --- 701 800 BG42284.12 (527) TCTGGCACCGGGTTTTGGCCATCCCAAATGGGAACAGGAGATCAATTGCTTCATTTTACAACCCATCAATGGAAGCCACCATAGCTCCTGCAGCACAGCT BG42288.1 (589) --- BG42289.1 (1) ---CCAAATGGGAACAGGAGATCAATTGCTTCATTTTACAACCCATCAATGGAAGCCACCATAGCTCCTGCAGCACAGCT 801 900 BG42284.12 (627) GGTGGATAAAGCAAACCAGGAGGTGGAACAAAGATATCCCAAGTTTGTGTTCGGAGACTACATGTCGGTTTATGCTGAACAGAAGTTCCTCCCAAAGGAA BG42288.1 (589) --- BG42289.1 (78) GGTGGATAAAGCAAACCAGGAGGTGGAACAAAGATATCCCAAGTTTGTGTTCGGAGACTACATGTCGGTTTATGCTGAACAGAAGTTCCTCCCAAAGGAA 901 930

BG42284.12 (727) CCAAGGTTCCAATCTGTGAGGGCAATGTGA BG42288.1 (589) --- BG42289.1 (178) CCAAGGTTCCAATCTGTGAGGGCAATGTGA

圖10. 阿拉伯芥AtACO5同源之苦瓜contigs核苷酸序列比對

Fig. 10. Alignment of Momordica charantia contigs which are homologus to AtACO5 nucleotide sequence. The gray boxes represent conserved nucleotides, respectively.

圖 11. 乙烯訊息傳遞途徑基因表現差異

Fig. 11. Representative gene expression profiles of the ethylene signal transduction pathway in Momordica charantia. The two color boxes represent gene expression level in control (C) and GA treated (T) bitter gourd. Log2 fold change was correlated to control. The color scale is shown at the bottom. Red and green colors indicate higher and lower expression, respectively. Gray color indicates that no transcript was detected in untreated samples. The ethylene signal transduction pathway was referenced to the pathway in Arabidopsis (Zhao and Guo, 2011). Gray words mean no contigs align to the reference genes. ETR1/2, ethylene response 1/2; ERS1, ethylene response sensor 1;

CTR1, constitutive triple response 1; EIN2, ethylene insensitive 2; ETP1, EIN2 targeting protein; EIN3, ethylene-insensitive3; EBF, EIN3-binding F box protein 1;

EIL, EIN3-like; ERF1, ethylene response factor 1.

圖 12. 類胡蘿蔔素及離層酸生合成途徑基因表現差異

Fig. 12. Representative gene expression profiles of the carotenoids and abscisic acid biosynthesis pathway in Momordica charantia. The two color boxes represent gene expression level in control (C) and GA treated (T) bitter gourd. Log2 fold change was correlated to control. The color scale is shown at the bottom. Red and green colors indicate higher and lower expression, respectively. The carotenoids and abscisic acid biosynthesis pathway was referenced to the pathway in Arabidopsis (Davison et al., 2002; Kim and DellaPenna, 2006; Seo and Koshiba, 2002). PSY, phytoene synthase;

PDS, phytoene desaturase; ZDS, zeta-carotene desaturase; CRTISO, carotenoid isomerase; LYC, lycopene cyclase; LUT1/2/5, lutein deficient 1/2/5; CHY, beta carotenoid hydroxylase; VDE, violaxanthin de-epoxidase; ZEP, zeaxanthin epoxidase;

ABA2/4, abscisic acid-deficient 2/4; AO, aldehyde oxidase; NCED, nine-cis-epoxycarotenoid dioxygenase.

圖 13. 離層酸訊息傳遞途徑基因表現差異

Fig. 13. Representative gene expression profiles of the abscisic acid signal transduction pathway in Momordica charantia. The two color boxes represent gene expression level in control (C) and GA treated (T) bitter gourd. Log2 fold change was correlated to control. The color scale is shown at the bottom. Red and green colors indicate higher and lower expression, respectively. Gray color indicates that no transcript was detected in GA3-treated samples. The abscisic acid signal transduction pathway was referenced to the pathway in Arabidopsis (Hubbard et al. 2010). PYR, pyrabactin resistance; PYL, pyrabactin resistance 1-like. PP2C; protein phosphatase 2C; HAI, highly ABA-induced PP2C gene; HAB, hypersensitive to ABA; ABI1/2, ABA insensitive 1/2; SnRK, SNF related kinase.

圖 14. 植物生長激素訊息傳遞途徑基因表現差異

Fig. 14. Representative gene expression profiles of the auxin signal transduction pathway in Momordica charantia. The two color boxes represent gene expression level in control (C) and GA treated (T) bitter gourd. Log2 fold change was correlated to control. The color scale is shown at the bottom. Red and green colors indicate higher and lower expression, respectively. The auxin signal transduction pathway was referenced to the pathway in Arabidopsis (Woodward, and Bartel, 2005; Tromas et al., 2013). Gray words mean no contigs align to the reference genes. TIR1, transport inhibitor response 1; AFB, auxin signaling F-box; ABP1, auxin binding protein 1; IAA, indole-3-acetic acid inducible; ARF, auxin response factor.

圖 15. 茉莉酸訊息傳遞途徑基因表現差異

Fig. 15. Representative gene expression profiles of the jasmonic acid signal transduction pathway in Momordica charantia. The two color boxes represent gene expression level in control (C) and GA treated (T) bitter gourd. Log2 fold change was correlated to control. The color scale is shown at the bottom. Red and green colors indicate higher and lower expression, respectively. The jasmonic acid signal transduction pathway was referenced to the pathway in Arabidopsis (Song et al., 2013).

Gray words mean no contigs align to the reference genes. JAR1, jasmonate resistant 1;

COI1, coronatine insensitive 1; JAZ, jasmonate-ZIM-domain protein.

圖 16. 雄蕊發育調控路徑基因表現差異

Fig. 16. Representative gene expression profiles of the stamen development pathway in Momordica charantia. The two color boxes represent gene expression level in control (C) and GA treated (T) bitter gourd. Log2 fold change was correlated to control.

The color scale is shown at the bottom. Red and green colors indicate higher and lower expression, respectively. Gray color indicates that no transcript was detected in untreated samples The stamen development pathway was referenced to the pathway in Arabidopsis (Cheng et al., 2009 ; Ito et al., 2007 ; Nagpal et al., 2005 ; Peng, 2009 ; Song et. al., 2011； Sun, 2011 ; Yu et al., 2004; Zhua et al., 2011). Gray words mean no contigs align to the reference genes. GID1, GA insensitive dwarf 1. GAI, GA insensitive. RGA, repressor or gai-3. RGL, RGA-like. LOX1, lipoxygenase 1. JAZ, jasmonate-ZIM-domain protein. MYB, MYB domain protein. AP3, APETALA3. PI, PISTILLATA. AG, AGAMOUS. ETR1, ethylene response 1. EIN3, ethylene-insensitive3. EIL, EIN3-like.

圖 17. 苦瓜花性決定之生物途徑示意圖

Fig. 17. Scheme of some of the biological pathways involved in Momordica charantia flower sex determination following GA application. Red boxes indicate genes that were up-regulated in GA treated sample compared with untreated sample, whereas blue boxes indicate genes that were down-regulated in GA treated sample compared with untreated sample. NCED, nine-cis-epoxycarotenoid dioxygenase; ACS, ACC synthase; LOX1, lipoxygenase 1; ARF, auxin response factor; MYB24, MYB domain protein 24.

圖 18. 以 RT-PCR 分析不同處理之苦瓜 ACC 合成酶基因的表現量

Fig. 18. The qRT-PCR analysis of the ACC synthase genes in the Momordica charantia of different treatments. The gene expression level in control sample was normalized to 1.

ddH2O, GA, ABA, NAA, KN and Eth means the 6-8-leaf stage bitter gourd seedlings which germinated from seed treated with double distilled water, 100 mg･L^-1 GA, ABA, NAA, KN and 25 mg･L^-1 ethrel at 5℃ for 15 days, respectively. Eth/sp and GA/sp means the bitter gourd seedlings foliar sprayed of 500 mg･L^-1 ethrel and100 mg･L^-1 GA at 6-8-leaf stage, respectively. The actin was used as the internal control gene. Each value represents the mean ± SE of three replicates.

圖 19. 以 RT-PCR 分析不同處理之苦瓜 ACC 氧化酶基因的表現量

Fig. 19. The qRT-PCR analysis of the ACC oxidase genes in the Momordica charantia of different treatments. The gene expression level in control sample was normalized to 1.

圖 20. 以 RT-PCR 分析不同處理之苦瓜花性相關基因的表現量

Fig. 20. The qRT-PCR analysis of the flower sexuality-related genes in the Momordica charantia of different treatments. The gene expression level in control sample was normalized to 1. ddH2O, GA, ABA, NAA, KN and Eth means the 6-8-leaf stage bitter gourd seedlings which germinated from seed treated with double distilled water, 100 mg

･L^-1 GA, ABA, NAA, KN and 25 mg･L^-1 ethrel at 5℃ for 15 days, respectively. Eth/sp and GA/sp means the bitter gourd seedlings foliar sprayed of 500 mg･L^-1 ethrel and100 mg･L^-1 GA at 6-8-leaf stage, respectively. The actin was used as the internal control gene.

Each value represents the mean ± SE of three replicates.

表 4. 外施激勃素於六至八葉期苦瓜對其花性之影響

Table 4. Effect of GA3 on number of male, female flowers and male to female ratio of Momordica charantia by foliar spray at 6-8 leaf stage.

Number of flowers

Male / Female ratio

Total Male Female

Control 858±78.5

837±77.8

22±0.7

37±2.3

GA

treatment 901±139.3

863±136.4

38±2.8

23±1.9

Data are the mean values ± SE of two replicates. Numbers within a row followed by different letters are significantly different by ANOVA, P=0.05.

表 5. 苦瓜轉錄體次世代定序概要

Table 5. Summary of next generation sequencing results of the Momordica charantia transcriptome.

Total reads 406,003,512 reads

Total reads length 41,006,354,712 nt

Filtered reads 401,960,352 reads

Filtered reads length 40,239,972,525 nt

De novo assembly 106,057 contigs

N

2,463 nt

表 6. 胡瓜花性相關基因於苦瓜之同源基因及表現情形

Table 6. List of Momordica charantia contigs homologous to cucumber sex-associated genes.

Name Contig ID FPKM Nearest

cucumber gene Description Control Treated McACO4.1

BG22617.1 640.85 607.91

CsACO1

Ethylene

CsERAF16

BG21294.1 2.29 2.55

CsERAF16

Ethylene-induced gene

CsERAF17

BG21769.1 0.57 1.58

CsERAF17

RNA was extracted for untreated control and GA treated seedling of bitter gourd sprayed by 100 mg･L^-1 GA for transcriptome analysis.

表 7. 本實驗室分離之苦瓜 cDNA 與其對應 contigs

Table 7. List of Momordica charantia cDNA isolated in our lab and their aligned contigs.

cDNA clone Gene Contig ID FPKM

pMcER287-4

McERS1

BG32732.1 46.19 44.32

pMCTR16-1

McCTR1.1

BG42151.1 4.27 4.89

AT5G03730 BG42151.2 15.86 19.24

pEIN2

McEIN2

BG26487.1 28.71 31.09

pMEIL324

McEIN3.2

BG21121.1 44.17 55.41

pMcEIL56

McEIN3.2

BG21121.1 44.17 55.41

pMcEDR111 AT1G18160

BG25544.1 15.84 18.23

BG25544.3 BG25544.5 BG25544.4

pMAEC28

McPIN1.1

BG35748.1 25.78 17.93

pMAEC43

McPIN1.1

BG35748.1 25.78 17.93

pMAEC93 AT1G70940 BG23381.1 13.36 15.79

AT2G01420 BG23385.1 52.65 66.39

pMAIC11

McAUX1.3

AT5G01240 BG31466.1 24.35 18.69

McAUX1.4

BG31472.1 86.26 65.81

pMcPAP-76 AT4G04020 BG12789.1 49.30 57.26

RNA was extracted for untreated control and GA treated seedling of bitter gourd sprayed by 100 mg･L^-1 GA for transcriptome analysis.

表 8. 苦瓜激勃素生合成途徑相關基因及表現情形

Table 8. Arabidopsis gibberellin biosynthesis-related genes and their orthologs in Momordica charantia.

Contig ID Gene ID FPKM

AGI Annotation

Control Treated

BG16911.1 McCPS1.1 0.65 0.44 AT4G02780

CPP SYNTHASE

BG17739.1 McKO1 12.51 12.96 AT5G25900 ENT-KAURENE OXIDASE 1 BG20689.1 McKAO1.1 13.89 14.96 AT1G05160

ENT-KAURENOIC ACID OXYDASE 1

BG17490.1 McKAO1.2 1.22 1.09 BG17490.2 McKAO1.3 0.11 0.07

BG38638.1 McKAO2.1 1.71 1.24 AT2G32440

ENT-KAURENOIC ACID HYDROXYLASE 2

BG38639.1 McKAO2.2 208.09 249.84 BG38639.3

BG38639.4

BG38636.1 McKAO2.3 44.35 51.53 BG13077.1 McKAO2.4 0.73 0.39

BG14156.1 McGA20ox2.1 0.21 0.48 AT5G51810

GIBBERELLIN 20 OXIDASE 2 BG9228.1 McGA20ox2.2 2.80 2.33

BG17029.1 McGA20ox2.3 1.07 0.39 BG6412.1 McGA20ox2.4 0.00 0.32

BG14156.1 McGA20ox3.1 0.21 0.48 AT5G07200

GIBBERELLIN 20-OXIDASE 3 BG53175.1 McGA20ox3.2 0.21 0.05

BG9447.1 McGA3ox2 2.19 3.13 AT1G80340 GIBBERELLIN 3-OXIDASE 2 BG33874.1 McGA2ox2.1 3.95 2.24 AT1G30040

GIBBERELLIN 2-OXIDASE 2 BG8254.1 McGA2ox2.2 0.06 0.24

BG19309.1 McGA2ox6 0.44 0.18 AT1G02400 GIBBERELLIN 2-OXIDASE 6 BG20162.1 McGA2ox8.1 0.06 0.21 AT4G21200

GIBBERELLIN 2-OXIDASE 8 BG20174.1 McGA2ox8.2 0.27 0.96

BG20174.5

BG49090.1 McGA2ox8.3 0.27 0.57 BG48466.1 McGA2ox8.4 0.22 0.30

表 9. 苦瓜激勃素訊息傳遞相關基因及表現情形

Table 9. Arabidopsis gibberellin biosynthesis-related genes and their orthologs in Momordica charantia.

Contig ID Gene ID FPKM

AGI Annotation

Control Treated

BG35789.1 McGID1B 5.26 9.25 AT3G63010

GA INSENSITIVE DWARF1B BG35789.2

BG16828.1 McGID1C 63.66 84.99 AT5G27320

GA INSENSITIVE DWARF1C BG16828.2

BG35379.1 McGAI.1 93.86 79.32 AT1G14920

GIBBERELLIC ACID INSENSITIVE BG36311.1 McGAI.2 14.74 9.37

BG19427.1 McGAI.3 60.52 53.96 BG16045.1 McGAI.4 77.33 99.63

BG16044.1 McRGA1.1 38.08 48.65 AT2G01570

REPRESSOR OF ga1-3 1 BG43062.1 McRGA1.2 0.63 1.89

BG19426.1 McRGL1.1 20.56 18.41 AT1G66350

RGA-LIKE 1 BG51724.1 McRGL1.2 1.63 0.98

BG16733.1

BG22846.1 McRGL2 1.29 0.82 AT3G03450 RGA-LIKE 2

表 10. 苦瓜乙烯生合成相關基因及表現情形

Table 10. Arabidopsis ethylene biosynthesis-related genes and their orthologs in Momordica charantia.

Contig ID Gene ID FPKM

AGI Annotation

Control Treated

BG18379.1 McSAM1.1 293.19 277.75 AT1G02500

S-ADENOSYLMETHIONINE SYNTHETASE 1

BG18379.2

BG18377.1 McSAM1.2 1151.13 1272.47 BG18377.2

BG8105.1 McSAM3 0.19 0.20 AT3G17390 S-ADENOSYLMETHIONINE SYNTHETASE 3

BG29132.1 McACS1 3.23 2.35 AT3G61510 ACC SYNTHASE 1 BG9684.1 McACS7 0.23 0.14 AT4G26200 ACC SYNTHASE 7 BG10149.1 McACS8.1 3.04 7.25 AT4G37770

ACC SYNTHASE 8

BG32424.1 McACS10 8.80 10.13 AT1G62960 ACC SYNTHASE 10 BG51419.1 McACS11 0.09 0.18 AT4G08040 ACC SYNTHASE 11 BG19736.1 McACS12 11.39 13.67 AT5G51690 ACC SYNTHASE 12 BG16781.1 McACO1 4.32 2.80 AT2G19590 ACC OXIDASE 1 BG22617.1 McACO4.1 640.85 607.91 AT1G05010

ACC OXIDASE 4 BG3185.1 McACO4.2 79.71 57.53

BG20621.1 McACO4.3 21.44 14.42

BG42284.12 McACO5 9.45 11.76 AT1G77330 ACC OXIDASE 5

表 11. 苦瓜乙烯訊息傳遞相關基因及表現情形

Table 11. Arabidopsis ethylene signal transduction-related genes and their orthologs in Momordica charantia.

Contig ID Gene ID FPKM

AGI Annotation

Control Treated

BG32732.1 McERS1 46.19 44.32 AT2G40940 ETHYLENE RESPONSE SENSOR 1 BG37469.1 McETR1 31.67 38.21 AT1G66340

ETHYLENE RESPONSE 1 BG37469.2

BG37469.4

BG18654.1 McETR2 32.88 40.70 AT3G23150 ETHYLENE RESPONSE 2 BG42151.1 McCTR1.1 4.27 4.89 AT5G03730

CONSTITUTIVE TRIPLE RESPONSE 1 BG25156.1 McCTR1.2 19.36 20.01

BG25156.3

BG25150.1 McCTR1.3 3.57 3.12

BG26487.1 McEIN2 28.71 31.09 AT5G03280 ETHYLENE INSENSITIVE 2 BG2025.1 McEBF1.1 71.35 91.88 AT2G25490

EIN3-BINDING F BOX PROTEIN 1 BG2584.1 McEBF1.2 74.53 85.50

BG7310.1 McEBF1.3 47.71 69.65 BG8936.1 McEBF1.4 0.48 0.66 BG49442.1

BG44947.1 McEBF2 0.00 0.29 AT5G25350 EIN3-BINDING F BOX PROTEIN 2 BG41424.1 McEIN3.1 142.14 189.49

ETHYLENE-INSENSITIVE3 BG41424.3

BG21121.1 McEIN3.2 44.17 55.41 AT3G20770 BG17837.1 McEIL3 4.87 5.13 AT1G73730

ETHYLENE-INSENSITIVE3-LIKE 3 BG17837.3

BG17837.4

BG35672.1 McERF1.1 1.17 1.76 AT3G23240

ETHYLENE RESPONSE FACTOR 1 BG41903.1 McERF1.2 1.26 1.24

BG1913.1 McERF1.3 6.17 11.18

表 12. 苦瓜類胡蘿蔔素生合成相關基因及表現情形

Table 12. Arabidopsis carotenoids biosynthesis-related genes and their orthologs in Momordica charantia.

BG35782.1 McPDS1 20.81 39.69 AT1G06570 PHYTOENE DESATURATION 1 BG28119.1 McPDS2.1 17.80 17.18 AT3G11945

PHYTOENE DESATURATION 2 BG28119.2

BG28119.3 BG28119.4

BG7864.1 McPDS2.2 1.14 0.98

BG11213.1 McZDS 48.15 41.15 AT3G04870 ZETA-CAROTENE DESATURASE BG25643.1 McCRTISO 21.31 17.18 AT1G06820 CAROTENOID ISOMERASE BG16858.1 McLYC.1 30.28 25.76 AT3G10230

LYCOPENE CYCLASE BG16858.2

BG19705.1 McLYC.2 1.02 0.93

BG4945.1 McLUT2.1 67.78 58.16 AT5G57030

LUTEIN DEFICIENT 2 BG54054.1 McLUT2.2 2.34 1.90

BG25287.1 McLUT1 25.32 22.52 AT3G53130 LUTEIN DEFICIENT 1 BG24857.1 McLUT5 38.29 25.02 AT1G31800 LUTEIN DEFICIENT 5 BG33129.1 McCHY1.1 18.52 25.34 AT4G25700

BETA CAROTENOID HYDROXYLASE 1 BG33129.5

BG22514.1 McCHY1.2 1.54 2.31 BG22514.2

BG24801.1 McZEP 15.26 13.91 AT5G67030 ZEAXANTHIN EPOXIDASE BG21689.1 McVDE1 68.08 67.29 AT1G08550 VIOLAXANTHIN DE-EPOXIDASE 1

表 13. 苦瓜離層酸生合成相關基因及表現情形

在文檔中苦瓜花性之轉錄體分析 (頁 32-72)