生物技术通报 ›› 2022, Vol. 38 ›› Issue (12): 184-193.doi: 10.13560/j.cnki.biotech.bull.1985.2022-0404
张业猛1,2,3(), 朱丽丽1,2,3, 陈志国2,3()
收稿日期:
2022-03-31
出版日期:
2022-12-26
发布日期:
2022-12-29
作者简介:
张业猛,男,博士研究生,研究方向:作物遗传育种;E-mail:基金资助:
ZHANG Ye-meng1,2,3(), ZHU Li-li1,2,3, CHEN Zhi-guo2,3()
Received:
2022-03-31
Published:
2022-12-26
Online:
2022-12-29
摘要:
探讨藜麦NHX基因家族生物学特性及盐胁迫下的表达模式,以期为藜麦抗盐品种选择提供理论依据。以藜麦含有Na+/H+ exchange结构域的蛋白质序列为基础,对其基因结构、蛋白质motif、理化性质以及盐胁迫下转录表达进行分析。结果表明,藜麦基因组中含有46个NHXs基因家族成员,氨基酸长度为109-1 820 aa,分子质量为11 472.74-204 381.38 Da,等电点为5.00-9.27,亲水性为-0.226-1.112,不稳定指数为21.71-50.73。进化树分析可以将其分为5个亚组,在进化树中位置相近的NHXs基因具有相近的结构。同一亚组的NHXs蛋白具有相似motif结构,与拟南芥、小麦和玉米为直系同源蛋白,且多数定位在细胞膜和液泡中。通过盐胁迫下藜麦转录组数据分析发现,随盐胁迫时间增加,其中有26个NHXs基因的转录水平提升,9个下降,11个无明显变化。此外,35个受到盐胁迫诱导表达的NHXs基因,通过GO注释发现,主要参与离子、细胞稳态和离子跨膜运输过程。NHXs基因结构和蛋白质motif的多样性,推测不同亚组NHXs在盐胁迫中参与不同的响应过程,其盐胁迫下不同的表达模式也表明在盐胁迫响应过程中发挥重要作用。
张业猛, 朱丽丽, 陈志国. 藜麦NHX基因家族鉴定及盐胁迫下表达分析[J]. 生物技术通报, 2022, 38(12): 184-193.
ZHANG Ye-meng, ZHU Li-li, CHEN Zhi-guo. Identification and Expression Analysis of NHX Gene Family in Quinoa Under Salt Stress[J]. Biotechnology Bulletin, 2022, 38(12): 184-193.
基因Gene | 上游引物Forward primer(5'-3') | 下游引物Reverse primer(5'-3') |
---|---|---|
AUR62041754 | ATGGAGGAGGGGAAGGGA | TTAAGAGGTACCCTTTTTTTCCT |
AUR62027477 | ATGTTCGATCGTGCTTTGGAG | TCATGAATCATGAGACCCCCCTT |
AUR62043892 | ATGGACCTTTCTGTGGTTGGC | CTAGTAACAAGCCCAAGGAAC |
AUR62017691 | ATGGAGGATCAGCAGATTTCTCC | TCAGTTACGAGTACTGTAGACTT |
ACT1[ | GTCCACAGGTGCTTCTAAG | AACAACTCCTCACCTTCTCATG |
表1 藜麦NHXs基因实时荧光定量引物
Table 1 RT-qPCR primers of NHXs in quinoa
基因Gene | 上游引物Forward primer(5'-3') | 下游引物Reverse primer(5'-3') |
---|---|---|
AUR62041754 | ATGGAGGAGGGGAAGGGA | TTAAGAGGTACCCTTTTTTTCCT |
AUR62027477 | ATGTTCGATCGTGCTTTGGAG | TCATGAATCATGAGACCCCCCTT |
AUR62043892 | ATGGACCTTTCTGTGGTTGGC | CTAGTAACAAGCCCAAGGAAC |
AUR62017691 | ATGGAGGATCAGCAGATTTCTCC | TCAGTTACGAGTACTGTAGACTT |
ACT1[ | GTCCACAGGTGCTTCTAAG | AACAACTCCTCACCTTCTCATG |
基因 ID Gene ID | 氨基酸 Number of amino acids/aa | 分子质量 Molecular weight/Da | 等电点 pI | 亲水性 Gravy | 不稳定指数 Instability index | 亚细胞预测 Predicted location |
---|---|---|---|---|---|---|
AUR62017691 | 514 | 57 092.24 | 5.38 | 0.439 | 50.73 | 液泡Vacuole |
AUR62044447 | 618 | 69 784.72 | 5.54 | 0.373 | 32.07 | 细胞膜、液泡Cell membrane,vacuole |
AUR62023671 | 510 | 55 482.83 | 7.06 | 0.747 | 26.34 | 细胞膜、液泡Cell membrane,vacuole |
AUR62022028 | 618 | 69 784.72 | 5.54 | 0.373 | 32.07 | 细胞膜、液泡Cell membrane,vacuole |
AUR62020843- | 517 | 58 012.47 | 8.38 | 0.460 | 32.20 | 细胞膜、液泡Cell membrane,vacuole |
AUR62017877 | 109 | 11 472.74 | 6.74 | 1.112 | 21.71 | 细胞膜、液泡Cell membrane,vacuole |
AUR62021694 | 665 | 74 913.30 | 6.68 | 0.403 | 30.77 | 液泡 Vacuole |
AUR62019781 | 804 | 86 860.99 | 8.88 | 0.360 | 40.84 | 细胞膜、液泡Cell membrane,vacuole |
AUR62021703 | 607 | 68 131.50 | 5.57 | 0.432 | 47.58 | 液泡 Vacuole |
AUR62008394 | 598 | 66 617.46 | 5.97 | 0.352 | 33.74 | 液泡 Vacuole |
AUR62021702 | 576 | 64 055.50 | 5.94 | 0.387 | 35.34 | 液泡 Vacuole |
AUR62022953 | 713 | 79 785.44 | 8.69 | 0.291 | 34.83 | 液泡 Vacuole |
AUR62009662 | 684 | 75 352.40 | 6.43 | 0.385 | 37.95 | 细胞膜、液泡Cell membrane,vacuole |
AUR62004319 | 1 227 | 131 988.87 | 5.08 | 0.050 | 41.71 | 细胞核 Nucleus |
AUR62027477 | 486 | 52 731.20 | 7.08 | 0.696 | 32.95 | 细胞膜、液泡Cell membrane,vacuole |
AUR62019779 | 801 | 86 091.87 | 8.44 | 0.360 | 36.40 | 细胞膜、液泡Cell membrane,vacuole |
AUR62043892 | 676 | 76 288.29 | 7.91 | 0.300 | 32.05 | 细胞膜、液泡Cell membrane,vacuole |
AUR62008393 | 607 | 67 909.32 | 5.65 | 0.440 | 47.78 | 液泡 Vacuole |
AUR62009709 | 867 | 96 471.46 | 9.23 | 0.297 | 46.97 | 细胞膜、液泡Cell membrane,vacuole |
AUR62033334 | 1 214 | 130 455.31 | 5.09 | 0.084 | 39.66 | 细胞核 Nucleus |
AUR62041754 | 349 | 36 309.11 | 8.89 | 0.781 | 25.43 | 细胞膜、液泡Cell membrane,vacuole |
AUR62033325 | 1 181 | 126 888.57 | 5.16 | 0.113 | 39.66 | 细胞核 Nucleus |
AUR62001874 | 429 | 47 260.98 | 6.45 | 0.636 | 33.44 | 细胞膜、液泡Cell membrane,vacuole |
AUR62001864 | 753 | 83 016.74 | 7.88 | 0.396 | 38.15 | 细胞膜、液泡Cell membrane,vacuole |
AUR62009707 | 704 | 79 132.21 | 8.20 | 0.242 | 41.03 | 细胞膜、液泡Cell membrane,vacuole |
AUR62000934 | 551 | 60 928.18 | 6.49 | 0.534 | 34.53 | 液泡 Vacuole |
AUR62027069 | 791 | 85 976.49 | 5.20 | 0.320 | 34.07 | 细胞膜、液泡Cell membrane,vacuole |
AUR62019780 | 801 | 86 024.90 | 8.13 | 0.387 | 40.33 | 细胞膜、液泡Cell membrane,vacuole |
AUR62036238 | 759 | 85 128.10 | 8.39 | 0.229 | 38.77 | 液泡 Vacuole |
AUR62018573 | 779 | 86 152.60 | 8.41 | 0.299 | 31.27 | 细胞膜、液泡Cell membrane,vacuole |
AUR62026204 | 778 | 86 638.91 | 7.90 | 0.174 | 38.40 | 液泡 Vacuole |
AUR62009395 | 1 646 | 182 928.70 | 8.87 | -0.226 | 39.57 | 液泡 Vacuole |
AUR62008401 | 665 | 74 930.13 | 6.52 | 0.407 | 31.41 | 液泡 Vacuole |
AUR62009673 | 1 820 | 204 381.58 | 6.58 | 0.332 | 37.00 | 细胞膜、液泡Cell membrane,vacuole |
AUR62001875 | 1 340 | 150 822.00 | 6.78 | 0.353 | 33.17 | 细胞膜、液泡Cell membrane,vacuole |
AUR62001876 | 674 | 75 685.34 | 5.54 | 0.340 | 39.06 | 细胞膜、液泡Cell membrane,vacuole |
AUR62022963 | 492 | 53 620.84 | 9.00 | 0.703 | 40.62 | 细胞膜、液泡Cell membrane,vacuole |
AUR62000862 | 544 | 60 147.10 | 6.08 | 0.470 | 32.40 | 液泡 Vacuole |
AUR62021295 | 548 | 59 615.20 | 5.58 | 0.675 | 25.39 | 细胞膜、液泡Cell membrane,vacuole |
AUR62030239 | 527 | 59 096.47 | 5.00 | 0.185 | 42.68 | 液泡 Vacuole |
AUR62003573 | 797 | 86 437.67 | 6.33 | 0.318 | 38.20 | 细胞膜、液泡Cell membrane,vacuole |
AUR62036236 | 844 | 93 703.46 | 9.27 | 0.309 | 48.18 | 细胞膜、液泡Cell membrane,vacuole |
AUR62003491 | 1 211 | 133 817.30 | 6.26 | 0.088 | 35.14 | 细胞膜 Cell membrane |
AUR62009672 | 648 | 71 771.52 | 5.28 | 0.331 | 33.94 | 细胞膜、液泡Cell membrane,vacuole |
AUR62026390 | 759 | 84 042.11 | 6.83 | 0.273 | 37.52 | 细胞膜、液泡Cell membrane,vacuole |
AUR62033999 | 491 | 53 035.55 | 7.08 | 0.709 | 31.97 | 细胞膜、液泡Cell membrane,vacuole |
表2 藜麦NHXs蛋白的理化性质及亚细胞定位
Table 2 Physicochemical properties and subcellular localizations of quinoa NHXs proteins
基因 ID Gene ID | 氨基酸 Number of amino acids/aa | 分子质量 Molecular weight/Da | 等电点 pI | 亲水性 Gravy | 不稳定指数 Instability index | 亚细胞预测 Predicted location |
---|---|---|---|---|---|---|
AUR62017691 | 514 | 57 092.24 | 5.38 | 0.439 | 50.73 | 液泡Vacuole |
AUR62044447 | 618 | 69 784.72 | 5.54 | 0.373 | 32.07 | 细胞膜、液泡Cell membrane,vacuole |
AUR62023671 | 510 | 55 482.83 | 7.06 | 0.747 | 26.34 | 细胞膜、液泡Cell membrane,vacuole |
AUR62022028 | 618 | 69 784.72 | 5.54 | 0.373 | 32.07 | 细胞膜、液泡Cell membrane,vacuole |
AUR62020843- | 517 | 58 012.47 | 8.38 | 0.460 | 32.20 | 细胞膜、液泡Cell membrane,vacuole |
AUR62017877 | 109 | 11 472.74 | 6.74 | 1.112 | 21.71 | 细胞膜、液泡Cell membrane,vacuole |
AUR62021694 | 665 | 74 913.30 | 6.68 | 0.403 | 30.77 | 液泡 Vacuole |
AUR62019781 | 804 | 86 860.99 | 8.88 | 0.360 | 40.84 | 细胞膜、液泡Cell membrane,vacuole |
AUR62021703 | 607 | 68 131.50 | 5.57 | 0.432 | 47.58 | 液泡 Vacuole |
AUR62008394 | 598 | 66 617.46 | 5.97 | 0.352 | 33.74 | 液泡 Vacuole |
AUR62021702 | 576 | 64 055.50 | 5.94 | 0.387 | 35.34 | 液泡 Vacuole |
AUR62022953 | 713 | 79 785.44 | 8.69 | 0.291 | 34.83 | 液泡 Vacuole |
AUR62009662 | 684 | 75 352.40 | 6.43 | 0.385 | 37.95 | 细胞膜、液泡Cell membrane,vacuole |
AUR62004319 | 1 227 | 131 988.87 | 5.08 | 0.050 | 41.71 | 细胞核 Nucleus |
AUR62027477 | 486 | 52 731.20 | 7.08 | 0.696 | 32.95 | 细胞膜、液泡Cell membrane,vacuole |
AUR62019779 | 801 | 86 091.87 | 8.44 | 0.360 | 36.40 | 细胞膜、液泡Cell membrane,vacuole |
AUR62043892 | 676 | 76 288.29 | 7.91 | 0.300 | 32.05 | 细胞膜、液泡Cell membrane,vacuole |
AUR62008393 | 607 | 67 909.32 | 5.65 | 0.440 | 47.78 | 液泡 Vacuole |
AUR62009709 | 867 | 96 471.46 | 9.23 | 0.297 | 46.97 | 细胞膜、液泡Cell membrane,vacuole |
AUR62033334 | 1 214 | 130 455.31 | 5.09 | 0.084 | 39.66 | 细胞核 Nucleus |
AUR62041754 | 349 | 36 309.11 | 8.89 | 0.781 | 25.43 | 细胞膜、液泡Cell membrane,vacuole |
AUR62033325 | 1 181 | 126 888.57 | 5.16 | 0.113 | 39.66 | 细胞核 Nucleus |
AUR62001874 | 429 | 47 260.98 | 6.45 | 0.636 | 33.44 | 细胞膜、液泡Cell membrane,vacuole |
AUR62001864 | 753 | 83 016.74 | 7.88 | 0.396 | 38.15 | 细胞膜、液泡Cell membrane,vacuole |
AUR62009707 | 704 | 79 132.21 | 8.20 | 0.242 | 41.03 | 细胞膜、液泡Cell membrane,vacuole |
AUR62000934 | 551 | 60 928.18 | 6.49 | 0.534 | 34.53 | 液泡 Vacuole |
AUR62027069 | 791 | 85 976.49 | 5.20 | 0.320 | 34.07 | 细胞膜、液泡Cell membrane,vacuole |
AUR62019780 | 801 | 86 024.90 | 8.13 | 0.387 | 40.33 | 细胞膜、液泡Cell membrane,vacuole |
AUR62036238 | 759 | 85 128.10 | 8.39 | 0.229 | 38.77 | 液泡 Vacuole |
AUR62018573 | 779 | 86 152.60 | 8.41 | 0.299 | 31.27 | 细胞膜、液泡Cell membrane,vacuole |
AUR62026204 | 778 | 86 638.91 | 7.90 | 0.174 | 38.40 | 液泡 Vacuole |
AUR62009395 | 1 646 | 182 928.70 | 8.87 | -0.226 | 39.57 | 液泡 Vacuole |
AUR62008401 | 665 | 74 930.13 | 6.52 | 0.407 | 31.41 | 液泡 Vacuole |
AUR62009673 | 1 820 | 204 381.58 | 6.58 | 0.332 | 37.00 | 细胞膜、液泡Cell membrane,vacuole |
AUR62001875 | 1 340 | 150 822.00 | 6.78 | 0.353 | 33.17 | 细胞膜、液泡Cell membrane,vacuole |
AUR62001876 | 674 | 75 685.34 | 5.54 | 0.340 | 39.06 | 细胞膜、液泡Cell membrane,vacuole |
AUR62022963 | 492 | 53 620.84 | 9.00 | 0.703 | 40.62 | 细胞膜、液泡Cell membrane,vacuole |
AUR62000862 | 544 | 60 147.10 | 6.08 | 0.470 | 32.40 | 液泡 Vacuole |
AUR62021295 | 548 | 59 615.20 | 5.58 | 0.675 | 25.39 | 细胞膜、液泡Cell membrane,vacuole |
AUR62030239 | 527 | 59 096.47 | 5.00 | 0.185 | 42.68 | 液泡 Vacuole |
AUR62003573 | 797 | 86 437.67 | 6.33 | 0.318 | 38.20 | 细胞膜、液泡Cell membrane,vacuole |
AUR62036236 | 844 | 93 703.46 | 9.27 | 0.309 | 48.18 | 细胞膜、液泡Cell membrane,vacuole |
AUR62003491 | 1 211 | 133 817.30 | 6.26 | 0.088 | 35.14 | 细胞膜 Cell membrane |
AUR62009672 | 648 | 71 771.52 | 5.28 | 0.331 | 33.94 | 细胞膜、液泡Cell membrane,vacuole |
AUR62026390 | 759 | 84 042.11 | 6.83 | 0.273 | 37.52 | 细胞膜、液泡Cell membrane,vacuole |
AUR62033999 | 491 | 53 035.55 | 7.08 | 0.709 | 31.97 | 细胞膜、液泡Cell membrane,vacuole |
图 4 拟南芥、小麦、玉米和藜麦的NHXs蛋白的系统进化分析 AT:拟南芥;Tr:小麦;Zm:玉米;AU:藜麦
Fig. 4 Phylogenetic analysis of NHXs proteins in Arabidopsis,wheat,maize and quinoa AT:Arabidopsis thaliana;Tr:Triticum aestivum L.;Zm:Zea mays;AU:Chenopodium quinoa Willd.
图5 藜麦NHXs基因表达模式与GO注释 A:NaCl胁迫下NHX基因家族表达模式;B:NHX基因家族GO注释;C:NHXs基因RT-qPCR分析
Fig. 5 Gene expression patterns and GO annotations of NHXs in quinoa A:Expression pattern analysis of NHX family under NaCl stress. B:GO annotation of NHX gene family. C:RT-qPCR analysis of NHXs genes
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