Molecular Characterization, Evolution and Expression Analysis of Ammonium Transporter from Four Closely Related Bactrocera Species (Tephritidae)
Abstract
:1. Introduction
2. Materials and Methods
2.1. Insect Rearing and Sample Collection
2.2. RNA Isolation and cDNA Synthesis
2.3. Homology Search and Molecular Cloning
2.4. Bioinformatic Analyses
2.5. The Phylogenetic Analysis of the Amt Family
2.6. Analysis of Expression Patterns of Amt in Different Peripheral Tissues
2.7. Statistical Analyses
3. Results
3.1. Identification and Sequence Characteristics of Amts
3.2. Phylogenetic Analysis of Amts
3.3. Structural Prediction of Amt and Analysis of Conserved Functional Sites
3.4. Expression Profiles of Amts in Different Peripheral Tissues
4. Discussion
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Conflicts of Interest
Appendix A
PCR Primers | PCR Conditions | |
---|---|---|
BdorAmt | BdorAmt-CDS-F: ATGACTTCAAAAGTGACAAATGTCT BdorAmt-CDS-R: TCAATCGATCCATGCAAAATTGATAT | 98 °C for 3 min; then 35 cycles of 98 °C for 10 s, annealing at 60 °C for 15 s, extension at 72 °C for 2 min; followed by a final extension of 72 °C for 10 min; stored at 10 °C |
BcorAmt | BcorAmt-CDS-F: ATGACTTCAAAAGTGACAAATGTCTC BcorAmt-CDS-R: TCAATCGATCCATGCAAAATTGATAT | 98 °C for 3 min; then 35 cycles of 98 °C for 10 s, annealing at 60 °C for 15 s, extension at 72 °C for 2 min; followed by a final extension of 72 °C for 10 min; stored at 10 °C |
BcucAmt | BcucAmt-CDS-F: ATGACTTCAAAAGTGACAAATGTCTCAG BcucAmt-CDS-R: TCAATCGATCCATGCGAAATTGGTAT | 98 °C for 3 min; then 35 cycles of 98 °C for 10 s, annealing at 60 °C for 15 s, extension at 72 °C for 2 min; followed by a final extension of 72 °C for 10 min; stored at 10 °C |
BtauAmt | BtauAmt-CDS-F: ATGACTTCAAAAGTGACAAATGTCT BtauAmt-CDS-R: TCAATCGATCCATGCGAAATTGGTAT | 98 °C for 3 min; then 35 cycles of 98 °C for 10 s, annealing at 60 °C for 15 s, extension at 72 °C for 2 min; followed by a final extension of 72 °C for 10 min; stored at 10 °C |
BdorAmt(q-PCR) | BdorAmt-q-F: GAACACCGGATTCGTCACT BdorAmt-q-R: CGCGTAGCTGTTCAATGCTC | 95 °C for 3 min; then 39 cycles of 95 °C for 10 s, annealing at 55 °C for 30 s, +Plate Read; followed Melt Curve 65 °C to 95 °C, increment 0.5 °C, for 5 s; Plate Read |
BcorAmt(q-PCR) | BcorAmt-q-F: CTTTGTGCTCTGGTGGGGAT BcorAmt-q-R: CATCATGACGCCAAAGCGAG | |
BcucAmt(q-PCR) | BcucAmt-q-F: CGCCATGATGGACGTATGGA BcucAmt-q-R: CCCCTAACCGGTCGAACAAA | |
BtauAmt(q-PCR) | BtauAmt-q-F: GCTCGGTTCACTGGTCTCAA BtauAmt-q-R: CCCCTAACCGGTCGAACAAA | |
BdorActin-2(q-PCR) | BdorActin-2-F: GTGTGATGGTTGGTATGGGA BdorActin-2-R: GGCTGGGGAGTTGAAGGTTT | |
Bcor_GAPDH2(q-PCR) | Bcor-F: CTTCAACTGGTGCCGCTAAG Bcor-R: ACCCTTCATTGGACCCTCTG | |
Bcuc_GAPDH2(q-PCR) | Bcuc-F: TGCTACTACTGCCACCCAAA Bcuc-R: ACGGAAAGCCATACCAGTCA | |
Btau_GAPDH2(q-PCR) | Btau-F: TGCTACTACTGCCACCCAAA Btau-R: ACGGAAAGCCATACCAGTCA |
Appendix B
Gene Name | Accession Number |
---|---|
Bactrocera neohumeralis Amt2 (BneoAmt2) | XP_050328977.1 |
Bactrocera tryoni Amt2 (BtryAmt2) | XP_039952923.1 |
Bactrocera latifrons Amt2 (BlatAmt2) | XP_018797628.1 |
Bactrocera oleae Amt2 (BoleAmt2) | XP_014087802.1 |
Anastrepha ludens Amt2 (AludAmt2) | XP_053956833.1 |
Ceratitis capitata Amt2 (CcapAmt2) | XP_004518340.1 |
Anastrepha obliqua Amt2 (AoblAmt2) | XP_054726403.1 |
Rhagoletis pomonella Amt2 (RpomAmt2) | XP_036333465.1 |
Musca domestica Amt2 (MdomAmt2) | XP_058984514.1 |
Teleopsis dalmanni Amt2 (TdalAmt2) | XP_037949734.1 |
Musca vetustissima Amt2 (MvetAmt2) | XP_061386255.1 |
Lucilia cuprina Amt3 (LcupAmt3) | KAI8129331.1 |
Drosophila melanogaster Amt (DmelAmt) | NP_001097800.1 |
Aedes aegypti Amt2 (AaegAmt2) | XP_021700161.1 |
Anopheles gambiae Amt2 (AgamAmt2) | XP_318439.4 |
Anopheles coluzzii Amt2 (AcolAmt2) | XP_040218851.2 |
Appendix C
Appendix D
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Gene Name | Blastx Best Hit (Reference/Name/Species) | E-Value | Score |
---|---|---|---|
BdorAmt | XP_049304235.1 putative ammonium transporter 2 [Bactrocera dorsalis] | 0 | 1211 |
BcorAmt | XP_049304235.1 putative ammonium transporter 2 [Bactrocera dorsalis] | 0 | 1204 |
BcucAmt | XP_054081809.1 putative ammonium transporter 2 [Zeugodacus cucurbitae] | 0 | 1209 |
BtauAmt | XP_054081809.1 putative ammonium transporter 2 [Zeugodacus cucurbitae] | 0 | 1097 |
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Zhang, J.; Wang, Q.; Liu, C.; Liu, J.; Qian, Q.; Ru, C.; Liu, L.; Yan, S.; Liu, W.; Wang, G. Molecular Characterization, Evolution and Expression Analysis of Ammonium Transporter from Four Closely Related Bactrocera Species (Tephritidae). Life 2024, 14, 1114. https://doi.org/10.3390/life14091114
Zhang J, Wang Q, Liu C, Liu J, Qian Q, Ru C, Liu L, Yan S, Liu W, Wang G. Molecular Characterization, Evolution and Expression Analysis of Ammonium Transporter from Four Closely Related Bactrocera Species (Tephritidae). Life. 2024; 14(9):1114. https://doi.org/10.3390/life14091114
Chicago/Turabian StyleZhang, Jie, Qi Wang, Chenhao Liu, Jiaying Liu, Qian Qian, Chuanjian Ru, Leyuan Liu, Shanchun Yan, Wei Liu, and Guirong Wang. 2024. "Molecular Characterization, Evolution and Expression Analysis of Ammonium Transporter from Four Closely Related Bactrocera Species (Tephritidae)" Life 14, no. 9: 1114. https://doi.org/10.3390/life14091114