Cloning and Spatiotemporal Expression Analysis of IGF1R Gene cDNA in Alopex lagopus (Arctic Fox)
Abstract
1. Introduction
2. Materials and Methods
2.1. Experimental Materials
2.1.1. Experimental Animals
2.1.2. Main Reagents
2.2. Experimental Methods
2.2.1. Tissue Sample Collection
2.2.2. Total RNA Extraction and cDNA Synthesis
2.2.3. Cloning, Sequencing, and Bioinformatics Analysis of IGF1R Gene CDS Region
2.2.4. Spatiotemporal Expression Analysis of IGF1R Gene in the Arctic Fox
3. Results and Analysis
3.1. IGF1R Gene CDS Amplification Results
3.2. Sequencing Results and Sequence Analysis
3.3. Similarity Analysis and Phylogenetic Tree Construction of IGF1R Gene
3.4. Structural and Functional Analysis of Arctic Fox IGF1R Protein
3.4.1. Physicochemical Properties and Hydrophobicity
3.4.2. Transmembrane Domain and Subcellular Localization
3.4.3. Phosphorylation Site Prediction
3.4.4. Secondary and Tertiary Structure Prediction
3.4.5. Protein Interaction Analysis
3.5. Tissue-Specific and Spatiotemporal Expression Patterns of IGF1R Gene in the Arctic Fox
4. Discussion
5. Conclusions
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
Abbreviations
IGFIR | Insulin-like growth factor 1 receptor |
cDNA | Complementary DNA |
IGF1 | Insulin like growth factor 1 |
IGF2 | Insulin like growth factor 2 |
MC1R | Melanocortin 1 receptor |
GAPDH | Glyceraldehyde-3-phosphate dehydrogenase |
CDS | Coding sequences |
ORF | Open reading frame |
DEPC | Diethylpyrocarbonate |
UTR | Untranslated regions |
PI | Isoelectric point |
LSM | Least squares method |
SNP | Single-nucleotide polymorphism |
FSH | Follicle-stimulating hormone |
GH | Growth hormone |
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Primers | Sequences | Annealing Temperature | Application |
---|---|---|---|
IGF1R-1 | F: GCAGAAGGAGCAGATGACATTC R: GAGGTGCGAGGACTGGATTAT | 54 | PCR amplification |
IGF1R-2 | F: AATCCAGTCCTCGCACCTC R: AGGAGGCTGTGGCTAGATTG | 60 | PCR amplification |
IGF1R-3 | F: GGAGTTCTAACAATCTAGCC R: ACCTGCGTGAATGTAAGAC | 40 | PCR amplification |
IGF1R | F: GTCCTTCGCTTCGTTATG R: CTTGTTCTCCTCGCTGTAG | 60 | qRT-PCR |
GAPDH | F: GGGTGATGCTGGTGCTG R: ACGGTCTTCTGGGTGGC | 60 | qRT-PCR |
Analysis Tools | Websites | Function Prediction |
---|---|---|
ExPASy | https://web.expasy.org/protscale/ (accessed on 24 February 2025) | Physicochemical properties |
ProtSca | https://web.expasy.org/protscale (accessed on 24 February 2025) | Hydrophilicity/hydrophobicity |
Novopro | https://www.novopro.cn/tools/tmhmm.html (accessed on 24 February 2025) | Transmembrane domain |
SMART | http://smart.embl-heidelberg.de/ (accessed on 24 February 2025) | Functional domain |
NetPhos-3.1 | https://services.healthtech.dtu.dk/services/NetPhos-3.1/ (accessed on 25 February 2025) | Phosphorylation sites |
SOPMA | https://npsa.lyon.inserm.fr/cgi-bin/ (accessed on 25 February 2025) | Secondary structure |
SWISS-MODEL | https://swissmodel.expasy.org/ (accessed on 25 February 2025) | Tertiary structure |
STRING | https://cn.string-db.org/ (accessed on 25 February 2025) | Protein–protein interaction |
Species | Similarity of Mature | Amino Acid Number | GenBank Accession No. |
---|---|---|---|
Canis familiaris | 99% | 538 | XP_545828 |
Sus scrofa | 98% | 538 | NP_999337 |
Equus caballus | 98% | 538 | XP_001489815 |
Bos Taurus | 97% | 538 | XP_606794 |
Homo sapiens | 97% | 538 | EAX02222 |
Macaca mulatta | 97% | 538 | XP_001100407 |
Rattus norvegicus | 93% | 538 | AAA41392 |
Mus musculus | 96% | 538 | NP_034643 |
Gallus gallus | 84% | 538 | NP_990363 |
Coturnix japonica | 84% | 538 | BAF73401 |
Xenopus laevis | 79% | 538 | CAA90517 |
Cyprinus carpio | 71% | 538 | AAN52152 |
Quail | Chick | Monkey | Human | Pig | Cattle | Horse | Fox | Dog | Rat | Mouse | Xenopus | |
---|---|---|---|---|---|---|---|---|---|---|---|---|
Chick | 0.006 | |||||||||||
Monkey | 0.134 | 0.132 | ||||||||||
Human | 0.137 | 0.134 | 0.002 | |||||||||
Pig | 0.128 | 0.125 | 0.008 | 0.010 | ||||||||
Cattle | 0.134 | 0.132 | 0.018 | 0.020 | 0.014 | |||||||
Horse | 0.134 | 0.132 | 0.012 | 0.014 | 0.008 | 0.016 | ||||||
Fox | 0.137 | 0.134 | 0.018 | 0.020 | 0.014 | 0.024 | 0.010 | |||||
Dog | 0.137 | 0.134 | 0.018 | 0.020 | 0.014 | 0.024 | 0.010 | 0.000 | ||||
Rat | 0.146 | 0.144 | 0.032 | 0.035 | 0.028 | 0.035 | 0.028 | 0.032 | 0.032 | |||
Mouse | 0.148 | 0.146 | 0.037 | 0.039 | 0.032 | 0.039 | 0.032 | 0.037 | 0.037 | 0.004 | ||
Xenopus | 0.165 | 0.157 | 0.203 | 0.205 | 0.198 | 0.205 | 0.203 | 0.205 | 0.205 | 0.208 | 0.210 | |
Carp | 0.248 | 0.240 | 0.276 | 0.274 | 0.271 | 0.271 | 0.274 | 0.271 | 0.271 | 0.287 | 0.284 | 0.266 |
Amino Acids | Number | Frequency (%) | Amino Acids | Number | Frequency (%) |
---|---|---|---|---|---|
Ala (A) | 31 | 5.8 | Lys (K) | 23 | 4.3 |
Arg (R) | 33 | 6.1 | Met (M) | 22 | 4.1 |
Asn (N) | 27 | 5.0 | Phe (F) | 20 | 3.7 |
Asp (D) | 24 | 4.5 | Pro (P) | 39 | 7.2 |
Cys (C) | 6 | 1.1 | Ser (S) | 40 | 7.4 |
Gln (Q) | 14 | 2.6 | Thr (T) | 17 | 3.2 |
Glu (E) | 49 | 9.1 | Trp (W) | 8 | 1.5 |
Gly (G) | 40 | 7.4 | Tyr (Y) | 22 | 4.1 |
His (H) | 8 | 1.5 | Val (V) | 41 | 7.6 |
Ile (I) | 25 | 4.6 | Pyl (O) | 0 | 0 |
Leu (L) | 49 | 9.1 | Sec (U) | 0 | 0 |
Histophysiology | GAPDH (Ct1) | IGF1R (Ct2) | Relative Expression (2Ct1-Ct2) | Brain Expression-Normalized (Set to 1) |
---|---|---|---|---|
Liver | 22.123 | 23.060 | 0.522 ± 0.109 | 1.906 ± 0.398 |
Spleen | 23.057 | 24.960 | 0.267 ± 0.085 | 0.976 ± 0.309 |
Kidney | 22.873 | 23.520 | 0.639 ± 0.230 | 2.331 ± 0.840 |
Lung | 19.745 | 22.150 | 0.189 ± 0.078 | 0.690 ± 0.286 |
Stomach | 20.323 | 26.160 | 0.018 ± 0.003 | 0.064 ± 0.011 |
Pancreas | 18.157 | 21.137 | 0.127 ± 0.107 | 0.463 ± 0.389 |
Brain | 19.767 | 21.633 | 0.274 ± 0.155 | 1.000 ± 0.565 |
Pituitary gland | 20.793 | 21.853 | 0.480 ± 0.154 | 1.750 ± 0.561 |
Testis | 22.333 | 20.883 | 2.732 ± 0.486 | 9.971 ± 1.773 |
Heart | 21.737 | 24.197 | 0.182 ± 0.052 | 0.663 ± 0.190 |
Muscle | 20.443 | 21.273 | 0.563 ± 0.057 | 2.053 ± 0.208 |
Histophysiology | GAPDH (Ct1) | IGF1R (Ct2) | Relative Expression (2Ct1-Ct2) | Muscle Expression in 4-Month-Old Male Foxes-Normalized (Set to 1) |
---|---|---|---|---|
2-month-old male fox liver | 22.030 | 26.527 | 0.044 ± 0.017 | 0.043 ± 0.016 |
4-month-old male fox liver | 18.353 | 19.450 | 0.467 ± 0.095 | 0.451 ± 0.090 |
6-month-old male fox liver | 22.123 | 23.060 | 0.522 ± 0.127 | 0.504 ± 0.120 |
2-month-old male fox testis | 21.120 | 19.760 | 2.567 ± 0.264 | 2.480 ± 0.250 |
4-month-old male fox testis | 21.107 | 20.020 | 2.124 ± 0.323 | 2.052 ± 0.306 |
6-month-old male fox testis | 22.333 | 20.883 | 2.732 ± 0.225 | 2.640 ± 0.213 |
2-month-old male fox muscle | 20.220 | 22.993 | 0.146 ± 0.008 | 0.141 ± 0.007 |
4-month-old male fox muscle | 18.460 | 18.410 | 1.035 ± 0.264 | 1.000 ± 0.250 |
6-month-old male fox muscle | 20.443 | 21.273 | 0.563 ± 0.066 | 0.544 ± 0.063 |
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Xu, W.; Fu, H.; Meng, X.; Sun, Y.; Ning, F.; Du, Z. Cloning and Spatiotemporal Expression Analysis of IGF1R Gene cDNA in Alopex lagopus (Arctic Fox). Life 2025, 15, 796. https://doi.org/10.3390/life15050796
Xu W, Fu H, Meng X, Sun Y, Ning F, Du Z. Cloning and Spatiotemporal Expression Analysis of IGF1R Gene cDNA in Alopex lagopus (Arctic Fox). Life. 2025; 15(5):796. https://doi.org/10.3390/life15050796
Chicago/Turabian StyleXu, Wei, Hualin Fu, Xiangyu Meng, Yiwen Sun, Fangyong Ning, and Zhiheng Du. 2025. "Cloning and Spatiotemporal Expression Analysis of IGF1R Gene cDNA in Alopex lagopus (Arctic Fox)" Life 15, no. 5: 796. https://doi.org/10.3390/life15050796
APA StyleXu, W., Fu, H., Meng, X., Sun, Y., Ning, F., & Du, Z. (2025). Cloning and Spatiotemporal Expression Analysis of IGF1R Gene cDNA in Alopex lagopus (Arctic Fox). Life, 15(5), 796. https://doi.org/10.3390/life15050796