CN113684281B - 一种采用snp分子标记技术鉴别润州凤头白鸭黑色喙性状的方法 - Google Patents
一种采用snp分子标记技术鉴别润州凤头白鸭黑色喙性状的方法 Download PDFInfo
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Abstract
本发明公开了一种采用SNP分子标记技术鉴别润州凤头白鸭黑色喙性状的方法,所述分子标记位于润州凤头白鸭基因组的11号染色体上,核苷酸序列如SEQ ID NO.1所示;所述分子标记位于第28397位点,突变碱基为A或T,同时利用该分子标记设计引物(SEQ ID NO.2和SEQ ID NO.3)进行PCR扩增检测。将所得扩增产物进行直接测序后判定受测个体基因型,可以确定润州凤头白鸭黑色喙这一性状,为有目的地对凤头白鸭喙色性状进行选择提供依据,提高润州凤头白鸭黑色喙的一致性。规范我国鸭遗传资源的生产经营,可以为润州凤头白鸭遗传资源保护提供有效保障,支持和引导鸭遗传资源监测评估。
Description
技术领域
本发明涉及一种鸭喙色的鉴定方法,具体是一种采用SNP分子标记技术鉴别润州凤头白鸭喙色的方法,属于畜牧学、分子生物学领域技术领域。
背景技术
对于畜禽而言,喙色性状是重要的品种特征之一。对于镇江润州地区发现的遗传资源群体--润州凤头白鸭而言,黑色喙则是其重要的外观特征。然而在长期利用传统的育种方法对该资源群体进行抢救性收集与整理中发现,经过多代常规的表型选育,后代喙色仍会发生性状分离,润州凤头白鸭群体的喙色表型始终未达到完全统一,严重影响群体的一致性。准确、高效的喙色选育方法,是润州凤头白鸭遗传资源保护亟待解决的关键问题之一。
SNP(Single Nucleotide Polymorphism,SNP),称为单核苷酸多态性,是第三代遗传标记,普遍地存在于所有物种基因组染色体上。研究发现,位于基因内部的SNP有可能直接影响蛋白质结构或表达水平,最终导致表型的差异,可作为重要分子标记,用于遗传资源的保护和重要经济性状的选育。
发明内容
本发明的目的是提供一种鉴定润州凤头鸭喙色的特异性分子标记,具体是一种采用SNP分子标记技术精确区分润州凤头白鸭喙色的方法,利用分子标记对鸭遗传资源进行特异性鉴定。
本发明的目的通过以下技术方案实现:一种采用SNP分子标记技术鉴别润州凤头白鸭黑色喙性状的方法,其特征是,所述分子标记位于润州凤头鸭基因组的11号染色体上,核苷酸序列如SEQ ID NO.1所示;所述分子标记位于第28397位点,突变碱基为A或T;鉴别时,包括以下步骤:
步骤1)、提取鸭的基因组DNA;
步骤2)、针对MITF-28397设计特异性扩增引物SEQ ID NO.2和SEQ ID NO.3,再以步骤1)的DNA为模板进行PCR扩增反应,获得PCR产物;
步骤3)、将步骤2)得到的PCR产物进行基因序列测序,通过基因测序序列和峰图判定鸭羽色基因型。
进一步的,所述步骤1)中待测鸭个体生物检材的基因组DNA为鸭翅下静脉血样基因组DNA。
进一步的,所述步骤2)中特异性扩增引物序列为:
MI-F1:TCCCCATTCTCCAAAGGTTAGC
MI-R1:TCCCCGTTGTCCCCACTTATTA
进一步的,统计步骤3)中获得的测序结果,判定鸭喙色的对应基因型:
若基因型为AA,表示受测个体为黑色喙纯合野生型基因型;
若基因型为AT,表示受测个体为黑色喙杂合基因型;
若基因型为TT,表示受测个体为黄色喙纯合基因型。
有益效果:本发明通过使用分子标记技术可以实现对润州凤头白鸭喙色性状的快速鉴定,操作简便且结果可靠,为润州凤头白鸭的遗传资源保护提供理论参考。同时为有目的的对润州凤头白鸭喙色性状进行选择提供科学依据,仅需一个世代的检测即可完成育种群在目标喙色性状(黑色)上的纯化。
附图说明
图1为鸭黑色喙和黄色喙个体基因型鉴定图。
具体实施方式
下面结合实施例、附图以及附图说明对本发明做进一步详细说明,所述是对本发明的解释而不是限定。
一种采用SNP分子标记技术鉴别润州凤头白鸭黑色喙性状的方法,具体步骤如下:
1.材料与方法;
1.1实验材料;
选择润州凤头白鸭黑色喙个体23只,黄色喙个体21只,其他鸭品种黑色喙个体59只,黄色喙个体70只。
1.2受测个体基因组DNA提取;
每只鸭个体于翅静脉处采血,用传统酚-氯仿法提取基因组总DNA,测DNA浓度和纯度,将DNA母液分装稀释至50ng/μl,于-20℃冰箱保存。
1.3以所提取基因组DNA为模板,通过PCR引物进行常规PCR反应;
1.3.1扩增引物
MI-F1:TCCCCATTCTCCAAAGGTTAGC,
MI-R1:TCCCCGTTGTCCCCACTTATTA;
1.3.2PCR反应体系及反应条件;
a.PCR具体反应体系见下表:
PCR反应体系
b.PCR具体反应条件见下表:
PCR反应条件
1.4将得到的PCR扩增产物送至擎科生物股份有限公司进行测序;
1.5使用ContigExpress软件针对测序结果进行统计分析,获得各群体基因型频率分布。
2.结果
根据图1判定待测个体在该位点的基因型。纯合黑色喙个体基因型为AA;杂合黑色喙个体基因型为AT;纯合黄色喙个体基因型为TT。
针对MITF-28397位点在不同喙色鸭个体的基因型进行统计分析(表1)。其中,在润州凤头白鸭黑色喙群体中发现52.17%纯合野生型基因型个体和47.83%杂合基因型个体(n=23),而在黄色喙群体中无纯合野生型个体,突变基因型和杂合基因型个体分别占61.90%和38.10%(n=21)。在其他鸭品种中,黑色喙群体发现69.49%的个体为纯合野生型,杂合基因型个体占30.51%(n=59);而在黄色喙鸭群体发现100%突变基因型(n=70)。
表1不同喙色类型(黑色、黄色)个体基因型
序列表
<110> 扬州大学
<120> 一种采用SNP分子标记技术鉴别润州凤头白鸭黑色喙性状的方法
<160> 3
<170> SIPOSequenceListing 1.0
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<211> 39457
<212> DNA
<213> 鸭(mallard)
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ctaacaagca tggtcggtgt cctccatgct cacactgctc cgcctgctac ttgttttcga 60
tgctccggga gacacggagg aaagtagcgg gtcagttact cctacgggag agagggtatc 120
gtccatcaag atatcttcca gtttggatcc catttttgta ggaacactgt aagtgccggt 180
cggttcagtc atgtttccga ggttgtcact gaaggtgatg gtaccgtcag tgagatcaag 240
ggtggtcgtg caagacaggt ctgtgtggtg cggcatgatg tcttggttgc agttgtccag 300
cgcgggttct tgtttgatga ccctgttgac catatcaggg gagcaaaggc cggtggatgg 360
cacaagggac agtccatgtg cccgagcttg catctcaagt tcctgtgaag gaggataaag 420
gagaaattaa aaataataaa aaaaaaatgg aagcaacatt tgtcatctta caagcttgtg 480
tattatgact tacagttctc ttctgttaga taggaaatgt ttcctccctt tccaatggtg 540
acaaacattt gcaccctgta cacttctacg ctgcagattt tcaaccaaat aacattctct 600
tgaagaagtg gataaatgct accaaacaca ggtccttcac agttctcaga acctacccag 660
acacgaaacc cagtgccctg cagaacagac atgttgtccc tcagactcct ttctcaaacc 720
agtgccaggg tagggaagct tggcgtcatg ttggggaagg aaagcatttc aaaagttgac 780
gaaaatgaaa cagcagaagt gaacacttag tgcttttcag aagttacgta tgaaagcaca 840
gtgtgtctat acaaggtagc tacttctttc tgagacactg caagggacat tgcacaccct 900
acagctctgg ttaggaggaa agtgaagata tcagcccaat ctgagcaaca ctactcctgc 960
aataaaaaca tacaggaagg ttacaggaag actttagaat catagaatca tagaatatcc 1020
tgagttggaa gggaccctta aggatcatca agtccaactc ttgacaccgc acaggtctac 1080
ccaaaagttc agaccatgtg actaagcgca cagtccaatc tcttcttaaa ttcagtcagg 1140
ctcggtgcag tgaccacttc cctggggagc ctgttccagt gtgcaaccac tctctctgtg 1200
aagaacttcc tcctgatgtc aagcctaaac ttcccctgcc tcagcttaac cccgttcccg 1260
cgggtcctgt cgctagtgtt aatagagaaa aggtctcctg cctctcgaca cccccttacg 1320
aggaagttgt agactgcgat gaggtctccc ctcagcctcc tcttctccag gctgaacagg 1380
cccagtgccc tcagccgttc ctcgtacgtc ttcccctcca ggcctttcac catcttcgta 1440
gccctcctct ggacactctc caacagtttc atgtcctttt tatactgtgg tgcccagaac 1500
tgcacacagt actcgaggtg aggtcgcacc agcgcagagt agagcgggac aatcacctcc 1560
ctcgacctgc tagcgatgcc gtgcttgatg caccccaggg tatggttggc cctcctggct 1620
gccagggcac actgctggct catattgaac ttgctgtcta ccacgaccct cttctaggct 1680
gctctccagt gtctcatcgc ccagtctgta cgtgcagcca gggtttcccc gtcccaggtg 1740
caggacccgg cacttgctct tattgaactt catgcggttg gcgatcgccc agctctccaa 1800
cctatccaga tccctctgca aggcctttcc accctcattc gagtccacaa ctcctccaag 1860
tttggtgtca tcagcaaact tgctcaaaat accttctatt cctacatcca gatcgtttat 1920
aaaaatattg aaaagtaccg gccctaaaat ggagccttga gggaccccac tagtgaccgc 1980
ccgccagcct gacgcagcgc cattcaccat aaccctttgg gccctgcccg ttagccaatt 2040
gctcacccat catatgatgt ttttatttag ctgtatggtg gacattttgt ccagtaggat 2100
cctatgggag accgtgtcaa aagccttgct gatgtccaaa aaaatcacat cagctggttt 2160
cccttggtcc accatacggg tgatcttatc ataaaaggaa atcaggttag ttaggcagga 2220
cctgcccttc acaaacccat gctggctggg accaatgact gctttgtccc ccaggtgcgc 2280
ctcaataagt tcgagaacca tcttctccat gattttacca ggcactgacg tgagactgac 2340
aggcctcact tcacatgcaa cttcacatgc aactagactt cacatgcaac taggattctt 2400
tgaatcttta ggaaaggctt aaatcccacc ttcacgttta atcgcaggca catttgactg 2460
aaggaagata ataaaagaat gcacttaaat gtcctgtttc aaatacagct gcaagacagg 2520
ttttttgctt ctacttatgc attatattaa gcaatcacaa tcactgttgt gattgcactg 2580
ttcctggata ttcttatcac ggaggttgat taacagcctc ttccatttcc aggctaagac 2640
aacaagcttg caatggcaca ctgcaacagc cacagagaag gtggaaaaga agcacttcca 2700
gcagctaatg tgccaaaatc agaatacaga ggaattcctt catcagaacc aacataaaaa 2760
gggaaaatac agagctcagg ttatttactg gcagatgtat tacccattat tccagggttg 2820
tttgtgtttt tttctcattg aaaagggagc cggagcagat gggtaatgca aaggaagatg 2880
aaagatgctt aaaaagcaat ctgacttcac tttgaaatac cactgcttga tgttgttctg 2940
cagaagggcc atgcttcttt atcttctcac ttgagcaagc agattgctca gcgaggtaca 3000
aaagctacgc tgaagctcat gccagcctgt tctattttaa atctgtttgc tgccagactt 3060
gactctcagg tatttcaagc ttcgttatct actctgtgga ttaactggtt cagggctctc 3120
ctgtatccag ccctgcgcac agcaacctgc agaagggctc agctgatcgc gctgcaagtg 3180
agcactaaca caagagtatc tgtggctcac agtccagcag catcgctgtc tttgtgttat 3240
gtgttcttac acccagttct acacatttct tgaattaacc ttttttgcag agtggtgtat 3300
atccagctat cctggaaaac agggttgata acaaaagctg tacaggcttc taggataagg 3360
ttacctatcc ggtttgaggt ggaggcaagg gattttatga gagaaatgaa gttcccaccc 3420
caaaactgct gtgtaataca gcctactggg aatctgggtg aagctaagaa tgcaacagcc 3480
catgatgaag acagaagaag gagggggaag atatattcat gccccccact gctccctcca 3540
gtgttgagga ggaaagctgt tctaatccta cagtctttaa ggaacagaga agaacaaatt 3600
acagctcctt gaacagaaag catcaacaac cacaaaaaca gccctacatc catagttcct 3660
ggttgtcctg cttcaaaaaa ctcttggggc tctcacattc aggctgcaac tcctctcaga 3720
ctcaaagcag actcaacttt taacacagcc gccaggaacc caatgctgca gtgacaacta 3780
tacttcccgc tgcccacttt gacctgtgag caatgtttac cctgcgaaca aaaaagcaga 3840
tgacagactc ctgcagaatg actcctgctg tcagagaaag gctcagatgt ggaagactcc 3900
agactgcgtt gcctctctga caggcagggc ctacattgtc ctgacagaat taaggaaacc 3960
cacaagcaaa acagattttg ttcattgcta ctttcagaaa cagcttcaga agaggacaca 4020
ccactacagc tccaagtcca caaaactccg caggcacaac taaagcacaa cagacagctg 4080
aactgaactt gattactatc aacaaataga tcatacgtga agtcctggtt ctggtctggc 4140
actcagaaaa aaatattact gtatagatct tttttatagt atggcttgca acttatgtta 4200
gctatcctag aacacaacca gaccagccac aagtgcttta acagaatact gaggagttta 4260
ggggccaggg aggagtgggc agaggagggc atctggttat tttttggtgt gattttattt 4320
attttaacta atgtatcctg ataattctgg aaaaattctg cagacatctc agtacagggt 4380
cactattaat taaaacagtg gctccaccta acccaaatcc ttataaagga ccttcagttg 4440
catctgtttg cagccacaca ctacaacatc aagagcactt gcaggttaag tatccacaaa 4500
cctgtattct gagcagcaga tgcctgttgg catgttccaa tttcttctgc ctgttctcaa 4560
gttcctttgt gcgttgctgt tctctttgaa gcttacgaat gtagtccact gatgctttca 4620
gaatagttcc cttattccag cgcatatccc ttcgaaatca acacaaaagg agaggttaag 4680
gctcccgtgg ggacttcacg ctgtagccag taccttcaga gcagcagctg tgattccacc 4740
tcacgctatg taacagccag ctaatacaat attcctccca agctgatgaa tgcagcaggc 4800
ttttgaaatg cactagaggg ggccagagca ctgctattgc aaggcagcga gaatcgtccc 4860
agctgaagga gagagcccag gcgctgctca gtggtttggg gctccccaca gccctctggg 4920
ccagagggat gagccccggc cttctgcaag cactgcaaag actgaagacc aagggctcct 4980
ttcattgcct tctgtgaggc tgcacaaaaa ccctacagga caccactcaa accatattac 5040
ttgctgtctg ggcgatgcct gatttcaggc agggtaatta aacgactgca ggggaactcg 5100
aattcataaa gaaccttata ggaattggtg tttctgtggg aggaagaccc gaggccgtgc 5160
agctcccaag cagcagcaag caacagtcct cctggtggca ctgtgccaca gcggggtccc 5220
agcaggaaga gcagagccca gcacagtgtg tttggatgct ttacgcttaa ctcttgctaa 5280
aaatataaat gctgtgttgc aaccgaggtc tgaccttcag tgaggggggt acagggctcc 5340
catattgccc ctcccacatt attcctacac cacacaaaag ggtgcctgca ttgtcttctc 5400
ccctgtgctg actttttttt ttatataagg agaaagttta gcacagagtg tacagaccaa 5460
agatccacct gttctgtagg ctgatgatgt gcaacattac cttgccttaa aggaagctaa 5520
agagcctctc taaatagtat ttattagtta aaataagaaa aagtctgccc caaagcctca 5580
ctgggcactg attcacattt aagtgacatt ttcaagctcc agagcaagag aggggttcat 5640
tgctatcgaa ccagcattag gtaaatgcca agacaaattg ataacactga aattacctaa 5700
aaagaactac aacccgcctt tctgggaaat taccgacaaa cacagcaaat tatcagttta 5760
acagacacag acctacagtc tttttttctt tcatatctct cttggaggga tggagaatgt 5820
gacggcttcc cagaggaact gccctccact tctacaaaaa gaaggatctg ttctgtcttg 5880
ccaagctatg acatctaata aattcaaagg atcgcttagc aagacaagca gctatgtagt 5940
cagctgagca gctggactag actatgcgtt cacatcctga cagctctctt gctggtacgg 6000
cggtcactca cagcatcagc cccaacgctg caaaagcaca gggccctacc aagatgctga 6060
gctgctgcca gggttctgtg actattttgt gacaaaagta atcaaattaa gaaccttcaa 6120
acacttctac tgccacagaa agccaacatc aaaagagaaa attcacgatt ttatgatttt 6180
atctccccta agatttcttt taaaaacaac caccaccacc ctcccgtcca cagactcctg 6240
ggcacatcat acttacgggt catttgactt gggtatcaaa gtgcctagtt cttttatacg 6300
atcgttaata ttgaatcttc ttcttcgttc aactttagga aaaacagcac agatgttaca 6360
agtcatatac acttaattac aaagtataaa gcttttaatt aaagatttct ttaagtgctt 6420
ttgaaattgt tgcctgaagc ttaaaatcta gtgacaatat agtaatttat tttgctcttc 6480
tacacataga gacagaagtt gtgtttacgc tttaaaaatg tgggactgtt gacgtggcaa 6540
gatctaaagt gagactctaa aaaatacgat acggctgtgt tagcacaaga ctccaataaa 6600
aataacgaaa agcgtttgtc ttccaagggg aaagtatcac aaaaatttgg agacaaaagt 6660
ttattgctca ggaaaatccc aaagcaggaa aagggacaaa aagtataggc atgtttatgt 6720
attctttaag attcaacact taagagtaca taaaaacaat caaacctaaa gatcaacttt 6780
cttgcattag cttttcatgt tattacaaaa gcttaccaaa aaggcctctg aatagcctaa 6840
ggcataaaag agcacctttt tttctattaa tacaaaagac tcttaagttg tcttttgata 6900
cttaaatcat aagatgctca ttgttctatt ttccttttct aaactcagaa atgagtgctc 6960
ccttcaaact ggaatttata aaattgtcac agtgaggatg tgcactggtg tttaaagtca 7020
taagctaact gcctacctta aattgtagtg gttttgttat gttttgtttt taaaaagcaa 7080
aaaatcacaa ggtgaatttg ttgaaagcaa ttggatggga tgttcccagt aattttactg 7140
gttaacacca cgtctgagcg catacttcca agaagtccag ataaacagat cgttacttgg 7200
aattagacca cccagaccat ttcacttttt tggaaaacgg tttaaaaatt gaagagtaga 7260
tttaagtaca cacttcagga gtatcagggg catttgatct atgggcataa cataatgaac 7320
cagtattaga cggagtggct tattcagatt taaaatgctc acattccaac acttaagaac 7380
aactatgatc attattaaaa ttggtgtaaa tagcaaagaa ccaaacttac tcaagttgtg 7440
attgtctttc ttttgcctct ccttagccaa tgctcttgct tctgactctg taggaaaaat 7500
acatgctgtg catgtgaatg aggtaattag aattaaagta attcaaacaa catttcagtt 7560
tgtgaaactt ttgtgaagga agcctgatca gcaactggca aacaaatcat tacagccttt 7620
ccgtaaagtg agaagggcct gtcacataca ttgtgtttac tgaaatccag agctggtatt 7680
caactttacc cctgcacctg cagagctttt gttcccacac ttcctatgct gacttctgtg 7740
atgagaacgt ttcccacgga ggccctgatt cagcaaggta tttgaggaca acaaattgct 7800
gaaatccaac atacctgctc ctacgtacac atgcagacaa catgcttatg taccacacag 7860
aggctgaacc agaggaacaa catgcttact gcattgctac ctgcacagga tgtgaagcag 7920
ccaggagtca gcagctccgc ttgctggaat tgagactatg tccttttgtg gcaacatgtg 7980
cacatcacca cagcactcaa ggtacggact taagtgacct actaaaaatg ctgtgttttc 8040
cagctatatt aattgttccc attatatcaa aaaagttttt aaaaaatcaa cacctttatt 8100
ggcaaatctc agatcataac tgttcttaac tttggtccac acatttatta ctcagacttg 8160
agcacgtata gtcaggcaaa cacataacta agcttgttat ttttcggttc tcatttgatc 8220
ttccatttcc caaactattt caactacttg tttgttctat ccattttaat ttctgaaaaa 8280
aaataaatga ggtgaaatcc ttaacccttt ttccctgttg cacttaaatt acaaaagcag 8340
caaacattag ttttgcagta gatcagtgag gacaagtgaa cctgtgatct attatcacac 8400
attcctttat gggaagaaaa cccttcactc aaacaatttt attttctgta ttattttctt 8460
tttatgcaat atggattatg tcccattcaa gccacagtaa ctggtgacga gggtttattt 8520
cccttggagt caccgtttgt gatagtcaaa ggcttggcca gatcaatgtc aaagtgactg 8580
ctgttaaatt tcagtttaat tcctaaactt aggacaagtt aaatgcaaaa cactttttct 8640
cctaatctat ggtagcagat aactaaattg cttgtttaca catgtgaagg tagcacatag 8700
atctgaacta agctaaatgt ctattttgac acactgagga cttctgtaag accacttcat 8760
tcactaattt cccctctttc tgccttcttt gcccaatgtc ttacacaatc acctaaagtg 8820
aatccaccag cctagacaca tggcaagttg agtgtttagc ttgcagtcta cagccatttg 8880
tagctttctg taactgaggc aatgaagctg tctgtaatgt aggaacctga gtagatctca 8940
tcacctccca ccctcctctt aacatagttc tacatgctgt tgaagttttt ctttctctag 9000
cagccactag tcattttgca ctttgtgtat cttcctttgg cgcagagctc aaattctgcg 9060
cactcataca aaatgcaaat atccaccagc tcagagacgt ggcatttcct ccagagaagc 9120
tcactcctgg gctattaaaa ctgctctact taaactaatt cctgtccaga acctgctttg 9180
ccactgagtc agcctgtgag ttttcacaac agaaagaaga gagacaagaa ataaactaca 9240
cctctaccag gtcagcacac agcatgactc tcagttgggc ttaagagcaa taatttctgc 9300
gtattaatat gattttgtca ctggttctag aatggatctc catttgttca tttgcagaca 9360
cctttgcttt gtgtatgtac cctacatgtg gttacatcct ttcatctaca agcgaagcaa 9420
aacagcaaaa aataaaacag gttttagaat atggagcttc ccttctaagc cttcaaacta 9480
tttagcttct gtatgcgtgt gcttgagagg gaaggttttt catgttccct ttcctgtccc 9540
ctaactccca cagcttattc agaggagacc aacttcaccg atggaagtac tgacatgaca 9600
agagcttcag aagatacttc attttcacct tgattcaaaa caaagacttt ttttttgtgc 9660
ctaattttac atttcatgga gtcacgatgt ccagatttct tattacaagt tatttgttca 9720
ttacagtcct cccctaatat taaagcctca acacgtagca ttcctcaacg tgaatcccag 9780
agagactcca gatgtcaagt tggtaaaatt taacgttgat agagcaaaca aggggtccaa 9840
actagcagct ctgcaggagt tgcatggaaa cttcctatgg cacaacattt tgaatgtcca 9900
gatttgactg caataattaa ggactttgcc ttaatttgtc acagtgaatg gacgcatgca 9960
aatacagctt attgccagct aatacatata ataataatta aaaaaaaaag actgcaaaaa 10020
gtagcaattc atcatttttt aaaaagaaat tgtaacagaa attatgtcca gtatcacctg 10080
aattcaaaac aattacacca tagctctgaa gtagtatttt agatgtattt acctgtgagc 10140
tcccttttta tattaggaag attagctgga cacgagttgc tgatgtttag tcctggaggc 10200
ggcatgcttt ggttgccata aaggtcaatc aaattcccag acacaggcaa ctagaaaaga 10260
caaggaagga gaaagatcag ttggtttgcc ttatgaaaaa gtttgtacag aacaaatgag 10320
ttatcacctg ctaggaagtc tgtgctcttt tccatgtgca aacagagctt gcagtaaggt 10380
cagcaaaaat tatgcatatg catgggcatg aggaataaac cacctacact ggttaagttc 10440
catcctctct tctcaggaag gtgcagacaa tactttatta ttattttttt gaaatgcaag 10500
gaagcacagc tccatttaag ttgaaaaaaa ttaagaaaga aaaggtaatg cacactgaca 10560
agtattgctg tattgttcct cacaattgct ctgccgaaag ctaagatctg taccctcagc 10620
aattccaagc aaaaaagcgc ctgaaagcgc ctacagaaga ctcatggagg atgcccccat 10680
ttttccaata acacgagtta aagttcttag caaatcatta caagcttgtc agaggacact 10740
gtattcatca atcacatttt tcatgtgatt tgaaggctag ttacagagag aacgttttca 10800
taagtttaac ttgtttttca ccttgcgtaa caaaaatatg caccggttgg aggaagactg 10860
ttttgacata catgtcctca attactcatt cattctgtta ttcttcttta ggtatgttta 10920
agcaaagaga ttcataagga aatagagtag tattccaaca agacttagca aagcaatccc 10980
ttaacattac aacgtaaatt taaagaaatt cccgtgtatc ctgagtggat tattttacct 11040
gcctattctt tgttttcaaa ctaggttttg agaagacagg tataaagtac tttgatacct 11100
tatctgagta gcaacatttc ttatctttgc tgcaaagaaa accagtccct ccaagacact 11160
gatcctgcag agattagacc attaacttca cacccacgta aataacgctg gggaacacaa 11220
tggagtgagt cacagcacgg gctattaaac tcctactgac agtccttgcc agactgagac 11280
aggagaaaat ttctctgcta cctagagcta ccataagaaa cgaagctgat aatgcacata 11340
ttgaagtgcc accacctaga cgccatgctg agatttgaca ggagacttct gaactgacag 11400
aggccaacag aaaaaggagg tcctctgact tggtcttcaa gtgacagagg aaaaaggacc 11460
tcaattctca gcaaatctag tcctacaatt tcctgcacac cacttcgctt ctgaatttca 11520
tatcaaaata tatattaaaa aaatagtaaa tctgttttag ttcccagctg tcctccacag 11580
agtgtgatcc tagttgcaac acacccacat taatgtcact gagaccatta tctcactgct 11640
ctacaaagcc gaccctcaga tcttttggcc cctcgacgca tccgtgacac aggagcagca 11700
gtggcgtcac acaaccttct gacttggacg tgctctttct attcatgctc ccaagtatgg 11760
tgagagcaca aaggacttgt ttgtaggcct gggacttgaa gttcagaacc agacatgtga 11820
aatctaatta ggaatttcag aggaaaaata aaaagatacc agaaactggc atctcaaatc 11880
ctgggtaaag cagctaggtt gtaacaaagc atggagaacg gtcgaggtca ccaagctgct 11940
ttaattattg caagataaac cctgcatgtt acgaaaggac tgtggattgt gattccacct 12000
ccctgtccac agatgtcaac atcatgttgg atttccatgt aaggttccgt aaggaacagt 12060
gcaatcacca caaaagggca ggggctgctg actgtgcaca ccgagtcctg caggaatggc 12120
ttgcaccaga agaacccacc tgagagctgg gcaacaaagc acaagcaatg ctgatgcatc 12180
actaactaaa gaaagatgaa gggagaaatt tgcatatctg attatctttt aaagtttgtt 12240
tctgtatttt aaatgttttc cttaaggctg tttgccgatt ttagctaaga catttaatac 12300
agcatctttc agatcatccc caagaaatta attcacctta actctctcta aaagtaaatt 12360
aagtaagagc agcttatgcc aggcctggtt tgtttccttc cttgagacag aaaggtgtgt 12420
ttttgttcac gagatttcat atcatgaaat atgatgggct ctaataaaat aagcactcac 12480
attcaggcca ttcaggctca gtgtgtatca ccgaggattc tgttccccag aaggtgctgt 12540
gctgctccct tacctacaag ggcacgctca ggggcagtcc ctgcaataag actcttctac 12600
tgctggtgtt cagcttccat tctgacacct tccagcacag cagcaaagct ggttaacttg 12660
tgacccagta ccaaagttta caaacatgtc cataggccct gtattgcaat gagccctttt 12720
caattaggag gtacttagaa acacccctcc gcttgttcta cactcaaatc tctgattttg 12780
tttatcaatt aagtactgtt accaaacgta acaaactggt cctttcattc tttgttggtt 12840
acggaacaat agcagccttt gttgtagtct cagcacagct atttataggt tgcgtctctt 12900
tagaaaccat ttcctctctt tatttttttt tccagctcta cttagctcag tctacagaca 12960
tcaattcctt attagtaaaa tactctgtgt tttcagagct aaatctcatt ccaaatgaaa 13020
agtgtttgtt atggcaggct cgtttttccc caaaccaatt gttgaggaac gtgtcttgtt 13080
ttccatcact tcacgactgg taccaggtta attcagaggc tgaaacatgc tttttgcatc 13140
gagcagttta atggtattag tagactgctc caagggcact atttaagggg ggaggttggt 13200
gggctgtttt ccgcagcatg caccatgcct ccaagagggc tgtgggaatc tgtgggagcc 13260
agactcggtt acatgacatc agacaagtac tgtcagcaaa tgtaaaccta tgctacaaaa 13320
atcgatgtta agtggtaagc tgcaaatttt ggcactagat aaaaaagaac gagacgccag 13380
cactggcaga catctactca gccagatcta acagcactta agcctaaggc aaaaaattca 13440
ctagagctac cctccacaaa agatcaaaat aagattatgc cagaagtacg attatgttaa 13500
tgctcgcagc aatgtgaacc agcaaacaga agtggcaact tggaggtaat gcgtgccttc 13560
ccttgctcac caactttccc atctcagcat tcttgtcctt tgtcccagac ttcctgggcg 13620
cccttctccc cattcagttt taagctttat tctttcacct ctggtttgct ttgaggggtt 13680
gctttttttc ctttcttttt tctaattttg taccatcttt gttgtcatct aacttgggac 13740
ttatctccat ttgcaatcag actaaatata tgatattaga gcttcacaat taaattagag 13800
cagctttggg cagggatgct aacacttaaa tgtggcttac agtttcttat ttgcacataa 13860
taagatggga acctggggtg ccaaaggttc cctctaggtt tccaaagcca agcttgatgc 13920
ccattaatat acccattaat atcaatgggt ataagcttac ttctgttagt tcaattcact 13980
cttctactat aatcttccca ctgattaacc tgaatttgat ttaaaaaaaa gcaaaattgt 14040
cctatgtatt ccctcaacct gactccaact tgcatttttg aactcacgtt cttcactgca 14100
attgtatact ttatgatctt caaagaaaag ctcattattc tccacagata gtgcgctaac 14160
tgcttaactg ctcagagaaa aaaaaaagtt acactccagt tgcacattca cttgaggagc 14220
agcgattagg cttaatgggg aaagcaaaag gaagggacca ttgtaccgtg tttgccatct 14280
gcaaggctgg gtccatcaag ccaaggattt cttcattgta gcttgattct agactaatta 14340
tgtcatcgat cacatcgtcc atctgcaata gtaaaggtga acaataaaaa atagaacagt 14400
tatcaaaacc aagaaggaac aaaggtctaa caaagcactt cagcaacaca ctggttctct 14460
gcagtatggg ccaaagcaga ggtctgcaca attctgccat actccaaatc taatccccct 14520
gacataactc tccctatcca gctagggctg tcagaccaca acaaagcttg aggcttcatg 14580
atctgttcca cacctcccat cagcagttac ctgcctacgg acagctgttc ctcccaggaa 14640
cagcagagga tatcggtgcc cgtacaacaa gcctcactaa ggatgtggaa ccctctacct 14700
tgttctgcac ctggtcccac taccttctca tctccaaaca cagggctcca gcactgcctg 14760
cccaccttcc accttgatca tccaggtgtg gggcaaaaag tgacttacag actgtaacgc 14820
ctgagccctg gcatctgcag agggaaggag aaggcttgct gctgaaaggc tgccctccct 14880
caacggggaa cctggtttat ttcctaatac tccatcccag gaagcaccac ccaagagtag 14940
ctcagtgatg attctaggtg gtgcaaacag tttttggggc tgggctggag ggttcctcca 15000
taagttccta catggtcaag gtgcaagaca gtcttccaaa gcactgctta gatgctctga 15060
gctgacctgc agcttgcacc tatttctttt tggatcacta taaaaaaata agcttaagtg 15120
acactaatca aaaataataa taatttcaaa aaaaacgtga aattgatagt acaaagagta 15180
gtttagggga ttttcatatc agcgcaataa agtatgatca gaatgaatta cactggctga 15240
aagtgagggc aagaaaggaa acaatagtta cagcccatat ctgaagtctt gttactaaaa 15300
taaaatgcag tctgatcaaa ataataataa taaaaaatac aaataattta aaaaatgact 15360
actgaggaca aaattaccat ggatggatta aagccttctc tctctacctc ccccttaagt 15420
gcaggtgtga tctgcctaaa aatgggctct tagaaactgt ctcagcagca acacgtacat 15480
gcagcttggg ggaaaaactg ctttaaaaac gacttcctga agtgagatcc taaaacaccc 15540
ttgataattt cttctgctat agtgttagag ttcttgttac agttctgaaa agggttaaca 15600
cttttctaga tgttttaatg cacttgatat caatggtaat atgcaaggac atggcagatt 15660
tgtaatgtaa gtgggggtta tgttgtttca tttcttaaat acattttaca ttcaacttgt 15720
taaatacatt actggtcaag tttactattt agcataagtt caaagaaaca ttctcattgg 15780
aaaggtgttc agtgctccaa ctcctgtgtc atccagtacc tgcatgcatg atgctcgaga 15840
gtgtgtattc agagccgggc actcactctc aaccctgctt tgctcttcaa atttataaaa 15900
tccctgcaga aataaaaagg aaaaaaaaaa agcaaagcat ggatctgagt tcatggcaca 15960
gtctcagtcc caaaaagagc acgattgaaa cagtgtcatg cacacatgtg cctgatgaca 16020
cagcaaagtg aagctggaaa taacttgaga aaaaaactgt ggtacaagtg agatgatagg 16080
atggaacttg gattttcact taatttaaaa ctatggagca caagtaattc ctcttgcatg 16140
tgtaaaagcc atttttaaat aactgttttt ggagttggat aaatttttcc tcacagagtg 16200
catctgtatt aaaattctgg cacaaaggct cagaggaagg atggggaaca aagggagctc 16260
tataattctc tcttgtttgc caagttgcta aaatattgac atgattcttt tacttgctaa 16320
tgtctttcaa ataaccttta ttttgcacct cagcacatac aacatccaac aaagtttttg 16380
ccagcaaaga tttaagacaa gattgtgaac gacttctagg catagaagcc tcaaatctgc 16440
agcgcagaaa cagcaaattg gaaacatttc aggatatact agctcagata atctggtgtt 16500
gaaactaagc aagagacttg ctcaaatttt agtccaaata tttctaaaac cctaaaatat 16560
ctaaaaaaaa aaagtctgtt gtttttcagt tatttttgtt atgctctttg ctctttctac 16620
actgaatggg ccataaataa cattattcca gaaagctaca gctaacaaaa ctgctttgta 16680
ttctccaaag gtcaactatt cactgcgtgc atcaagcaag caaaaaaaaa aatagaagtt 16740
ttaataagca taaactagct aaaatagttc aaaaaaggaa attacatgac agaggggttt 16800
ttttgtttgt tttttgccct aatgttttat acaaatacat attcatcagg caggcaaaga 16860
agcattaatt gcattttcaa actgagcagt tcagaaaaga agataaacgc aacatcttaa 16920
ttagcaaata tagtttgaaa cggaaagaaa gcagaatgca attctgtttc actgaagagt 16980
atgcacaaca agcgtgtatt acctcttttt cacagttgga gttaagcgtg agcatggcca 17040
tcgggctgtt gggcgcgctg cttccagttc caggcggcat gacatgatca ccgggctggt 17100
ttggacatgg caagctcagg gcttggttgg cgtgtttatt tgctagagtg gtagagaggt 17160
actgctttac ctgctgccgc tgggcttgct ggatatggta cttggttggg ttctcaaggt 17220
gagtctgcac ctgcatgaca ggagttagga aagggcttta ttgttttaca acgtcattac 17280
gtgtggcatc gatctgagct atacatctgt gacacagacc agctaagcat ttgactcaaa 17340
gttacaaacc cgtgcggctc caaggaactc aacagggctt cactaattca cgttcagtgg 17400
caaggaatga gttcctgaga gaatgttcca ataactgact tattaacagg aaagcaacga 17460
gttccacttc tcttccacat ggtgagattt cccacttaaa atcaatgctc cagttttcca 17520
gttcacacag acactctgca acaggtactt ctctgttcag accttcagca acacagtgtt 17580
taattttcac caaaatatgg agaaataagt gccacagcag gactcgtttt acttgtcaat 17640
agctcctaat aaaagctaaa tagtatcaac tgctatcctc tatcagattt caagtgaagg 17700
acagatttaa tgatttgttg ttggtttttc ttttgctttt taaaatatca aatgtgttat 17760
taaaaaaaaa aaaaaagtaa tgtcccattt tttgtctcta gatgatttat aactcagata 17820
tatggcctaa gtcacctaca aataagacag catgtctaaa aaaacacgtt acacaaagaa 17880
gatgcaagaa aagcagcgtc tccagtcaga tcaatcactg tcctcaatag cagccatttt 17940
tgtatttaag ataacctttc cagcacttca aaatattttc acatataaaa tatgaaaaaa 18000
agcacaccaa gctattctct tccactcgaa ataagcacca gaagaaaagt tttttttgtt 18060
tttttttttt ttaatggaaa aagtcaccat atcttggaaa tgccgtcacc tttctttgat 18120
cattaatatc tttatcaaaa ggtagaaaac aaacaagtaa aataatactc tgaaaagttt 18180
gtttgttttg aacagttcca gcaaagtgac ttctattacc gttacaactg accgtgcacg 18240
ttccccactc atttctgtgg aagaaattaa agcaaactag atacaagagg aaaaggattt 18300
tctgtccact gtaagccttc ttagcaagag cttttaaaat aagtctgcaa ataactcacc 18360
tgataatgat tatactcaag catttccagc ataatactta taggttgtga gagaagtgac 18420
tgcaatatcc tttgctttat gctactccag tctgagcgat tcacatatgg aagcaccata 18480
aacaagctgc cccaaaccag cagcagaaac tggaacacca ctgctcagac ttatagtaac 18540
ctccgctaat tagctacgca tgtagagaaa caaacagttc acacaggggc tgtgcaagtt 18600
tagccctaag tattggtatt ttttccctgg ctttgacgtc atgctttttc ataaacataa 18660
aaggaccttt taagtgatgg cctatcaaag tcagattgac agtttgctca aggctaactc 18720
gctattcatc tttcgttcca gtcatactga tagacaatgg tatttttctt tagcagtaag 18780
gttaaaagtt tttaagtacg gaggaagaaa taaccaaacg aatgccctac gagcccaacg 18840
cacatccacc agccatgggg cgctggatga gcccgcggcg ctatgcagcc ctggtgcagc 18900
cgggagccac agcagcccat ggccctgccc ttggcacccg ctgctcaggg cgccagctgc 18960
acccaggacc tgcagctcca cagggaggat ttgaggattt caaggagccg tgctgcttct 19020
tcagcctcct ttgtgcccat tcagtctgtc ccagctttca gcagcaaggc ttccagtgta 19080
ggaagggttg acaatgggct aaagcacatt gcaaatatac ctctgtccac ccaacaccat 19140
tactgcacac agtccctaca cctcccagac ctttcttatc cttgctactc aatcgcttga 19200
cagaagtgtt gtgctgaagc acatgcatct gaagttgctc tacttgtgct gttttccaaa 19260
gtctgctgca cagagaaaca tacttcagat agggtgttaa gagagcttaa cctgatttgg 19320
tgcaccaagt cccaacacag tccggcagcc tgctgcgcta agtgcagcac ttgctttgtg 19380
agtcccaagc ggtgttatcg attttcatat caccagtggt aagagttctg ctacccagat 19440
aaacctattt ttcacttaaa ttttcatctt ccaagtagat gaagttatca ttaaatgagt 19500
tcactttcgt aacccttatt ttaaaaacaa tcttacagta ttcgcacagt ccttttcctt 19560
ccccagcatc aaaaaccact catgctccct cagccagaat gagggacgga actaagcctc 19620
ctccctgcat caaccacatc aaccacagat ccatgctccc aggagaagga caagcaaaat 19680
tccaggatcc acatctccac tggcacaaaa ctgctcccca ggctcgctgt gcaagtgctc 19740
tgcgctgtgc cacaggtccc tgcttcgccc agtgcacaca cgcatgcctt ggcctcttcc 19800
ttctcactgc taaggtcagg catcaagact tctgtattag aaatggctaa agagtaaatc 19860
agcagtaaca aagcttacaa tgaaaggcaa agttgggatc ccgcacactg cctgatggaa 19920
cagatccgtg ctgatgtatt tgtctgccgt ttgcctgtaa ggactcccgg ccctattgtc 19980
tgaacctgca ggctgatgat acaagagctt gcaaaatgtg caagtatcat cacaatttac 20040
agaggtacta ataccccgaa ctgactccac gacacagaaa gttttgaaac acatgctgcc 20100
tacctgacat cagactcttg ttggctaaac aaacactaga aaacaaaaca tcttaccact 20160
attttttttt gtcttatttt gctactccat aatacttttt ttctttaata ggctgtgccg 20220
gcctttgttg cagggaaggc aagaaaaaaa aatgtgaaat tcagcttcag aaagaacaca 20280
gctcctttta aattccagca gacattaggg tatcatttaa taacagcatg acaaataaaa 20340
tcctgtgtct aagtagacag aagaagagca acaagagcat gaaaatgcag cgtttttact 20400
tcagctggaa catatttggc agccaactgg cacttctgac aggactttta gtcagaaact 20460
aggaagtaag gctggacatt tagcttctgt gttaggcttt tactgcctaa cagccctgca 20520
gagccctctc aagaatcgca tcaacctttt ccacctctgc ctctgaagtt atttcagttc 20580
tcttatatca gatttctacc aattacagaa gtgtaactat tctctgatcg ttttaacctt 20640
actgacaagt gtgttctctt tggcacaggg caacagactt caccaagagt ccagaggcta 20700
tttgggtgac gggaattcct cctcttcaca gggaacataa ctgctagaca aacagcactg 20760
aagatttaca tgcagcagaa ctgcaaagac aaattgtagt ctagctggtt taattcctaa 20820
cacaccacac agtcaccaca gtcactttgg ttcagaaaca acggcacagt cttgtggaag 20880
tattttgaac aaactttttt ttttttaatc ttgttataca ctggtaacaa ctagctctga 20940
ggaaatgcat tttctgtcag gtgtctggac ttcaaataag cagcagagaa gactattgct 21000
tttacagaga aaactatcag aagagctgtg gtcattttct gctagttgta cgcagttctc 21060
cagggccatg ccaccttctc acagaagtcc agtagtgaac tgcagaacac agaactgcct 21120
tttctgtcat taaattgctc tgaaatgatg caaaggggag atttggtgag agaacatttc 21180
ctctcagtct gtagctgatc caacatactt ccgtgtttga ctaaaagaac gaatgattaa 21240
tctcagaacg agggggcctg caacactggt attccagctc ctcacagcag tgcaacaaaa 21300
ccagcagatc ccacaggatg gaaagcatct ccacacggtt caggcagata tcacacctcc 21360
accactctta gtggtggggc acacaccgag ctgtgcctgc accgaagccc agcagcacaa 21420
caccccgtgc ttccctccta ggccatggtc aggcctagtt cttcaagcag gagctagcgt 21480
atggcgagag cacaggccct gctggtcaga acagcttacc ccggtttcct cctccaattt 21540
gataggaatg tatcccccaa gaggatacag cacccacttc tgaggaaaaa gcggattatt 21600
aaccctccaa acaaggagtt ggccagcaaa gctgaagagg ccccagtcct caattgtggt 21660
ggtacatctg caagagtctc ctgataacct atcatgatgc cgacagctct agtcccccac 21720
aggcagggaa ggagcacatt tgagttgcct gaatcacaca gaagattctt ctcatcaaaa 21780
aacaccactg aacctttctg tgaaagcaca gccaaaccac ttaatgcttc atttatgcag 21840
agacaggctt gggcacagga acataccgaa gatttagaga ggacagctaa ccaaagtggg 21900
aggaagaatt tttcccctca cccccacatt tttttttatt tttatttttt ttttaattaa 21960
aaagaccatt tgtttcaagt aaactgcctt aggacttaca gtttataatt acaaataaaa 22020
tacttttcgg tcggttttct cagttcaata gaactttttg ttgcttagac agctaaatgt 22080
catgggattt ttaagaaagg ttaaaattct gcgtcgttta acaattaatg gtacccaaag 22140
aaaaattata taaaaccaca tccagtgcca tgttttgcaa gagctacttt gctggcttca 22200
gttaggttcc ctgcaccagg aagccagctc agtatgctag cccaggcctt cacagctagt 22260
cttgttttta aagctgcttc aacccatcac tttcagggat tagcagcaag ttgttttcta 22320
ggtccttgaa tgaagtcaaa aacctgtaca aacaggtgtt tatcatttca catggggtaa 22380
agtctgagaa caactaaaag gaaagacaat caattactgc ctcttatatc aagatgactg 22440
cattttactg tcaaagcaca tatacattgt taacttgatt ttgagcagct ggcaaattag 22500
tcttaaacat aaataaccat tcaaagggga ttcttgtagt gtgagttatc attattaccc 22560
agccttaaaa gcctttattc tccctaggct ttctaatcca aattcaatat tctgcaaaca 22620
gtattcatca cagactggcc tattatgcct agaaatctga agcaccacat ataattccaa 22680
taattccaat tatttcatta aaatattgat catgggtttc actgcaatgc caggaagaac 22740
cctcacagta tccaaatcag agaaatttaa atatccagga agacaatggt ttaactcatt 22800
ctcaaagaca ttttcccaca tgcagcattc aaatgttaag cactgacatc acatttttcc 22860
ccttaaggac atctgtggtc tgaagtagct tcacattgta agttcagtgt tcttgctttc 22920
ttaaaccttt tcagtcctcc aggatgaaac agttacctca cttgaaatat aatcaaaaag 22980
aagttgattg ggttgcagta cgtatcattc ccaagggtag ctgggaattc agaaacagat 23040
atacctattt cataacttct caggctgttg ttctcccctt ctctccatct ttgctagagc 23100
atatttgctc cctctctttt agtccataac ccaggggcag tgactagaac aacactgacc 23160
agaaagccca gggagaaagc tgaaaagtta cttttactga ctggggaatg atgtgcaaag 23220
aagaatttat ttcagtattt tatcttagga catagccaaa gcaaggaaat cacatctgtt 23280
aacacttaat aaaactctct ggttacttac ttcatccaaa acaccctctg aacatgaaaa 23340
tcaaggcagg tacaagaaag ccatttaaga gtcagcagta tgattgcagt ctaagtggtt 23400
ccaacagtca tcttcaaaca cagttatgaa aaggcacatt gtacagaaag aacatcgttt 23460
acacaactga gaattgtttt gcccatcacc aaaaactcct ctgtgtaacg tgacattttc 23520
ctccaaggga aagtcatgga agcagacaaa aaaaagaaca aagtttttgt gattggtatt 23580
taaaaattgt gctttagcta caggagcaca tgatcagagt atctaaagaa attaaagaga 23640
ggttaaccaa tttgcttaag atgacaggaa ttggtgtaat ttacaggtta gtggcctgct 23700
agctgccatt ccgtaagtgt ggttttcgga cagaatctca tcctttgaac tgctatggtt 23760
ttctgttagg aaataaacat ttcagctgat tattttctga catacatgcc ataatttctc 23820
agttttttat agttactggg taggtaccag aaaccttttc aacatttcag gacagacaag 23880
taattgactc catgaaatgc cagacacaaa tctttgctac gatttcacct ataccagtca 23940
taaatatttt atgactggca gacaacacta ttggccacaa aactgtatct aaccagcaag 24000
cttgaattac tgccaaggta aaaaaaaaaa aacttgattc aagcacctca aatctacatc 24060
ctcctttctc atctgcacta gaggccttga cactgctcac attagggcat gaaagagaag 24120
actctcccat acaggaactg tcaatgacag acataaagct gctccataag atctctcgta 24180
agctggcaga gacacgtcag ctccagagga agcagaattg taggatcaga catctcagtt 24240
gccccaaggc ctgtactgat ggtctgaaac cacttctgtt cccagcttct ccttggaaag 24300
cctttcatag gaaaagagct atcatccata cttcctacag cagcaccaac ttaagattta 24360
tctagaccac tctaacacac ttcgctattg acttgaaaga gtcacttcag attggtaaaa 24420
atatttgtcc aaccttatct tcctcccaag gctgccaagc acggcattag ctagtgccag 24480
ctttgacctg cagtcacata tttagagcct gagcaagcct atcagttaca tttcacagga 24540
gttaccaatt ccgccaccct tccagcactc agtaacccct gctagacctg cacaacagaa 24600
catttccctt ccaggtccct ggttcactgc acctctgctg taaaacctgg tggcttcgaa 24660
cagccttcat tctacagcat ttctgatggc aagaaagaaa tgcagaacag caacactcat 24720
tctcaggagg gccttcacag gtcagcttac gacttaaagt acatcaactt tcacacgtac 24780
aaagggttta gagatttggg atggctttct agagcccaca gctgggagag catgtactta 24840
gctgtgcaaa aggaggaaca tgtatgtcct caaatactgc agggctctga ggcagtttcc 24900
acaggtgaga agaggcacct ttctccaggc cctggtttgc tagggtcact cactgtcaat 24960
cagctgccaa acatgacaga ttttaatcca gtaaatctgc tttgtccctg tatgggcttg 25020
tcttcacatg tatcatgcaa cactgacttt tcacaagtgg aatttaagct aaaactggcc 25080
attttgctca acctgtagat gaaagcaagc aaaacaaacc aaacaaacat tattttcata 25140
acgtttaaaa ctccctaccc tccacaccgg tgttgtcact acaaggtttg tgaaagcttg 25200
ctacatttta tagcatatga gatgaagtta gagcccagat aactgctggt gtagaggaac 25260
actatcttca gcacttgggg gccaagcaag gactaaggct gatgtcagat cgttcaaatt 25320
ctcagctcta gcttgtaaga aagaagtggt gttcagaata gtgcctgtgg gccatccaag 25380
tatttactta caaaatctca acattcagcc aaggagtgca agataggaag agatttaaag 25440
atggcactag atcaaaatat attgaacagc caaaagccac aaacagaacc atacttgtgt 25500
ccttcccttt tttgaacaga agttatttgt aagtgcaaat acaactccta gattattata 25560
ttcaagagga aaaaacctac tttggagagg cacactgcgc accaacagga catccacact 25620
tgttgcatac atatgttaaa aatgatgcag gtgaaatact gctacactct ttgttcacat 25680
aatccacggg gactgtgaga ctggcagcat gctccctcgg tccctgacaa aacacatacc 25740
aagacatgcc tctgctcgta agtgaagtct gaaaagagaa aacacgaacg ctaaacccaa 25800
cgaacaccac agtgcatgct gtgtttggcc tgtaccacta actggaagac cttcctacca 25860
tagcttttaa aggaatgccc aaaaataaaa gtcataattt tgaaaggacc atttatttta 25920
atgaatcact tagcactaca ctggctacag cacaaaggac acttgtttaa gtacgcgcct 25980
gtttggaggc tgttttctga cggacacgta gcgccatgag aaaaaattag caacagaatg 26040
aagatcttgt taattcaaaa cagttttatc cattaaaatg ccctgagctt ctaaatgtca 26100
cagttaaaaa aaaaattaga gacactttgc tgggctctat aatattttgc ctacagccta 26160
cgggtgtctc atgtcagccc caataggaaa ggtctggaga agcagaacaa agcagtccag 26220
atgcacccgt tttatccaca ctgctaatat gatcattaga aacagcctaa aataatacta 26280
cactgctgag ttcttttaag tgaaggaaat gcaagctggc ttaccaccga actgcttgat 26340
ttttatcctt ctgttcaacg agctgtgatg ttatttatga ctgtattaaa acctactgag 26400
ttgtagtaag gcagtttcag tacctcgcag cgctcgatca tcgtgctcac taatgtaaga 26460
caacccaagt caccttcatg caaactcaac cagctacaat ctgaagcatt ctgctaaata 26520
ccagaattgg gattttgttt ttggtacttt ggcaagccac aaaggtgcgt acagccctta 26580
cacagaggca agcctccgcc ctggaggaga attaacccag ccagccgaat tgatgccaca 26640
gagcatcaaa actgagaact gtgctgtcag cattactgct gacaggtgat ggcttttcct 26700
agcaagcaga gtgcttatgt ccctgacact tccattaagc aacattacat attttatttc 26760
ctgacatctc actcacaaag aaggtgttag agcagacatc acagtagcca caagagatta 26820
aaggtagcag cagtaagaaa ttaagtatgt ttattgcact catcttaaac cacaaggaaa 26880
tttagtgttt tagttgtttc ttggctacag gaatgctgcc ctaacacttt tgtaacacca 26940
agatgtattt aaggcaaaga gccataatgt tgtgtgacct cacctttaca ccccaggatc 27000
ttattcccgt aacagccaga tggtggctat cttctggctc caataaggaa aaaaaaaaaa 27060
aaaaaaaaaa aaaaaggcaa aagcacacac aagcttatcg taggctcatc tgtcaaagaa 27120
agatggcaac atttcagccc tttaaagatc tcaagcccta catacaccaa gatacagcat 27180
cacgatctat tttataccga caaaaacaac atccagccac tcggccactg ctgtaacctt 27240
cggcacacct gcccctccgt gtctcgcaga tgagctgttg aaaacccttc tcctcatgct 27300
atttcacata tggcaggcaa ataatcagtt acactgaaac aagaagcacc tgtccaccta 27360
aatgaccctt gacaggccag aagatggagc atttgcagca ccatcacctg aactggggca 27420
agctttccac acaaattcag tcagctgtgg ttggctctgc tcagccagcc gtaaccaaaa 27480
aacgcagtca ggatcattac ttccaagaga tgttgcacaa gcaattttta tttttatttt 27540
ttattttttt ttaaatcatg aatacttaaa agagaggctg tgaaaaaaat gctatatgaa 27600
aagctgaccc aatcttctct tttccagcac aaacccatgc cctggtagca gcagcctttt 27660
cagcactgac tctgtacaag agcaagggcc tgtgagctct cgatgctctg ctggctgcag 27720
tcctactgac acagcagggt taggactctc ccgacgggac cctgaatcgc attttgtggt 27780
gtggcagcac actaattcca ccttccgaaa cattttacag aaagctcttt tcaggggttt 27840
catctgaact tcaaaaaagc acaagtcaac agctcaccag gagaggcact tcacgagaca 27900
caaagccggg tgtgtgttat catccccatt ctccaaaggt tagctgcaga tcagtattgc 27960
caagaaattc agaagtattg gttttgtcag aagatcaaaa cgtctcttgc gagagacatt 28020
tagggcctcc aggcttggtc tcagaagaat cctattttaa accacactga tgtaaccctt 28080
caattgctgc tacaagaacg gaactaggtt cccaagggga aacaggcaca gcactttcca 28140
gcacagcctt actcgtgggg aaaatacaca caaagcctct tgctgctgga aaacaggcaa 28200
tgctgcatgt gtatcacgac acctgtcacc actgatggat agcgtacaca aagtttcagc 28260
ttcctaagag agtatggaag cataacccag ctccaggcaa gccgagtctg gacaattaga 28320
tggaggtaga gattagatgt ggagtaacta gatcctaact caatctcccc accttcctta 28380
ccccttttta tagctcaagc ataatcccat tttagcaaat cattatttta catgatggaa 28440
tgaacagctt gagatcttaa tgctttgaaa atgcaaatag gcacacaccg aactgtgttg 28500
gattttaaag gaaagtttaa atggaggaac aaagcaaggg agctggaggc agcagaccta 28560
atgacagcca gcatcgaggc ccccccatct cggtgtgtgc ccaccagcag tgtgctgcag 28620
gacacggggt gccaccaggg actggctgca gacaccacgt atgccccagg ccaccgctgc 28680
cagcccggct caggaccgat ctcacaggct gagttctcca tagtggctgc agagcagctt 28740
cttgtttaat ttcctctgac atgaatgtaa ttagtactca tcaacagcac tctgcgaaaa 28800
taaactcact tgtaataagt ggggacaacg gggaattgac ttccaaagca cgttgctgct 28860
ccacgctcca ggctgctggc caccagggct gcaaggaaca ctctcagcca cctctgctgc 28920
aaacagtcca tgtaacacag aactgaaccc cccaaaaacg ggtgtaaata agcaataggt 28980
tgaataatag cctgttttgc ttctgcctta ccccatctgg tggcctactg cttatgtaca 29040
gctggaagca acagttaatt gaggcgcgtt atcgggaatt gcagaactag agaaataaac 29100
gtggcactgg atgagctgcg gtgccctaac agttctctgc tgctgttccc tgagtgccat 29160
ggcagtcgca ctcctcactg catcccaggt ggagcatctc gatgccgtgc actcacacca 29220
cgtggagcca tcgggaccgg gcgccgtcac cgcagccccg cgagctggtc gtggcggcct 29280
caaccgaagc ggataagtgg aattaatacc aactcctgga gaaggtggaa actcagaaag 29340
actttctaag gatgaggctt gggagctgcc tgactcaggg agaagccttg gctggaagct 29400
gtcccctcgt tccttacatg aaatgtcgca gagagaggca gaaggcagct ctgagcaacg 29460
cacatgggaa gcatcgcttc gcacagagca cccacttaat ttaataagaa gcactccaga 29520
gcagaaggca ccacttttct tccctcaaaa agccaatata cattagaaaa gcataaattc 29580
ttttgaaaag ccacagtagc atgattaaat actatttttc cttttacaca acaaacctta 29640
cagggctgca cccttaagga atatatgaac tttcatgtga aaggagccca ttaactgctg 29700
cacaatgaat ggtgcctgtg gtccaagatt catttccttt tagaaagcag aaaggacgga 29760
gagcttacaa ctgctgcaca caactgctga ctcattcaag gtcactttac ttcttccttg 29820
aacaaactgt ttctttgaga cccagcctat cgtggtattt aagactgtag tttcaagagg 29880
gagtcaggga acacaatttt gttaggctgt gccaaatttg caggacccaa tattatttca 29940
aatttaagtc cgattttttt ttttcttaaa gtactattca gaaacgaaaa tccattttgg 30000
cagggtgggg agaggggatt aaatttcaca gaggtcactt taaacaagcc tacaagtact 30060
tccaaagccc ttaaaagtag attttataca gatttgtgtc atgtcagcat ggctatcaaa 30120
cagatttcaa gagacataat ttccataaaa cagtcagtag tactaggtta gttctagaaa 30180
tagtactaga ttatgtaaat aaagactttc aattattgag ttacgataga gatgactggc 30240
gtgtttttta aacttaagtc tcacttctaa gaatttgctg tttaacaaga caagaacgaa 30300
cttcaaagcc ctttctgaaa ggacagcgag caccacatgg cttcagcaac acagccccag 30360
atcgcacggg gctgcagcag cagtaagaga agctctgccc cactggcttt gaacagaggc 30420
tttccacttg ctacctgctc atctcttcct ctaaaaatca gccatgaagt ctgtaaagtt 30480
ttgatggttt cagctaggaa tttccaggtg acttttcgga agaaagcctg aaacagaacc 30540
tttgatttat ccaccgtgac acacaagctt cctaacaaat tcctagctca cttctgattc 30600
ggagtcagaa taatttctgc tttttccacc acaagtgcac tgagcacagc agtcgactgc 30660
aaggccttta actaggaaac aaaaaatgtc agtgttagat agggagggga aaaaacactc 30720
ccaaaatcac ggtgaggatt ttctagagct tacagcaatg actccaagtc ctttaaagat 30780
gcttgggtgc ccagcaccat ctgagacccc aagccctcct agcaaggcca cacagctccc 30840
tcccgatagc ctatgatgct ttttggggag cagatctgcc tctccccatg ttcagaagga 30900
aaagagagat agatacgaat acagtgcccc gggacagaaa aaatactctt taaaacacgg 30960
ctttttgttt gtgtttttta aacacaagac gcatttctcc tcatttcagt gtgcttgttt 31020
ttggatacat tgcagattcc aattttgatt ttctttattt aaagcattcc agctgcactt 31080
gctgaacaag tttactactt ccacttgctg ccagtagcag gtttactttc aagagagagc 31140
ctgccttcca cagattttct agcagcaatt tgtttccaca tgtaggaacc gttgtgtagg 31200
gcgcacaacc ggttaatggc ttaaagacac ctctgggaca tgagttggtg gaaaccaaat 31260
acaatataaa caagacaatt agaatcatgt gtgaaaatca gtttctatca cttgactgtt 31320
agccaccgga gaagggaacg gtctggaaag gcagcttcag cagcaactga gagctttccc 31380
aagaaataga cgctcttgga aaccataatg ccatctgctg agagattatg atagtgcaat 31440
gcaaggaatg acagccctga attagttctc cagtgcttac agcatttcat ctgctagctt 31500
aattactgta atatcgatgt tgaaaggcat gctgcttctg gtcatgttca aaaataaaaa 31560
gaaattgttt aaatgggata agttatccaa gggttttcaa agaactacct gagacatcac 31620
agcctactga taatcagcaa gtcaaggaca ctaaatcaca gaggtccata attgaaatgt 31680
tgttccagta tgctgaaagg atacaagtag gatgtgggac acacatattc atttggctgg 31740
gacactctaa cattgcaaaa acactgggaa ttaaatagct ctttttgatc tattatgttt 31800
ttaaagttac tctattttgt gctccgtatc tttacaagct tgagaatttg tacataattc 31860
taattcaact acaaaaatat atcaccaagt ctcaaaatta caaataggtt aagtgtgaaa 31920
ctggaatcca atagaagtgc cagcgacagc ctgcaggacc gagctgttac cccaatatcc 31980
caccagcact ctccatagaa atgtttggag gcagcaaggt atttttgtag ttggtagcgt 32040
agttgctgtg aactaaagat tggcaggata atgattccac aaagaccacc agatggatta 32100
tttcatttcc caatctgcca agcatctaaa atcaggccct gttttgctgg gtcacaacaa 32160
aacttaatag aacatgagga acaagaacca gccattggga tgcaaagggg atgcacgcat 32220
gacaagccag gaggatgttt ttccttattt gcactaaacc aagtctgccc ctgaaggtac 32280
tgatgaatta caagaggttc agaaagcctt ctgaaggatt caagatcagg cacgccccag 32340
attcaagatc taagcagaac actagcacag tctgcttttc ctctagccca caagtttcag 32400
gagtgttgca atctgagcct gtcattaatt attaaaggaa gatttacagt aataataatt 32460
taattctcta agtcagacag acagaactaa ccagatccac tggctagtaa ctgatggtgt 32520
atgaaatcct aagtggatgc tgccgcacag aactgttctt ccagagaagt gcaaaggcag 32580
cagcctggtg atccagttct agtctgaccc tctcatttca gttctgagga atgcctctcc 32640
tatgtgtaaa gaaagggaaa aaagattttt ctataatatt ttaactgaag aatgctgcat 32700
gagtgtcttc ctaaggaaac acaaaccaaa tgtgttccat cctcagctca gggacaattt 32760
ccactgcaag cagtacagag ctgcattact aaaatggatc cttttggtca aagaacaaga 32820
catggggctg gcacctatct gtgaggaata aagagttaaa acagctacca cttcctcaac 32880
tggcccactg ctattgtgag ggcagtgttc agctgaattc catcaggcca cgctgtctta 32940
acataacctg acgtgttcct cttcaagatc tcggaatttt tttctcctct ctgcttttca 33000
gcagcataca ctagtaaata aatacacccc aaaagccaaa tggtaatcca agtaaaaata 33060
atatttaatt atatagtggg gggtagtgag gggactgagt aacttttttt tttttttttt 33120
ttttttaaag taaaagactc gccaggtcct tcacagacaa cacaatactg gtttttgttt 33180
caagttcata acttggctct taaagtagtt gataatgaac cagcaggcat tattttaaat 33240
caaccagttt tattaaatga gccagaaagg aaagaggcag ggctaattac cagcattcct 33300
tgatgaatta cctagcggtg tgtggtaaat ctctgggcgt ctttgtcttt atccccaaat 33360
ttatccctta cttatttctc aagaggttaa aggacacggg gctaattaag gcaccaatcc 33420
aatgacagac agaatcattc aaggtaaggc tggtttttac tctttcaagc ttttgcatct 33480
ctgcctggta cttcttaatc taatttcttt tttttttttc cccccttaat ataactgctg 33540
gggaggggga ggaaaagcgc taatcttgtc acggtcaggc aggttgaaat tgccatgcag 33600
tgctgggcaa aatgctgcat atttgtaatt tagccaataa gaaattcctt aagcctctcc 33660
tgaatggcag gaatgtctct agcttgtcac ttccaattca gtcactgcgc aggttcggcg 33720
tctcgcagaa taatgctgcc aaactagctg aggaactgaa atatttgcga gtgtacataa 33780
tgcctaaaga ttactttaag cagatctgag ttttcacatt tcaacattta acgtctgtca 33840
aagctgtcga cagaggcctc cccagacctc cacatgactc cccccagcct tcagcgccgt 33900
ggggggctcc aggctgtgct gcagaggcaa ccagcacaac cccaggcacc agcactgccc 33960
ctttctgtgt tcttggctca accggtgcag ggtggtaagg tggttttatt ttaagctgac 34020
ttgggctgaa acaggatttg atacaaccta ggcaacacac acactgacac ccatagcaaa 34080
aggagatcgg ggatgaaggg tgaaagcagg ctttgcactc gcagtcaagc agtcagacac 34140
ttgtgcacag aagtacattg cccccttgaa ttccacgcat gtttaacgtc taataacctc 34200
tgtgttgagc aagagtttac attttacctc tagcactcac cacagacatc agcaccagtt 34260
ctcattctgg aggagcttac aacccactaa gcaagcaaaa gactgaaggc ctgagagata 34320
tcttacttac gtgaaggctt aggacaggct ttagaggttc tgtggttaaa agcaagtttc 34380
tccttctgaa gaatttgctg tgggctgaag ttacactcac cgactctgtg ttgtatctgc 34440
agttcagcaa gacctggtct ggcaaatcac ttctccgagt tacaactgca aatacatgcc 34500
aggctggctt aacagactgg cacacaccgc tagctgctat tgtcaggttc gattaaatta 34560
acttttgatc agataagcat ttccacaaac acacatttac tgtaacagat gctaccacac 34620
ctttatctcg acatcccgtt gttcctgttc tacagaactg taggacagat agattgatct 34680
cccatacaat gaagtctcgt ggtaattatt acaccaggaa tactatttct aactgcacag 34740
ccttccctgg tgagcagaag gactacatta acaccacagg tcgctctttg taacagagca 34800
cgctggaaaa ggtctgggag gaggcatttt ttgatacttg tccatagtca ttaacatcac 34860
tcagaggcat gcttggcatt tcaaaaccga agaaatacag ggtatctgat gcaaagagct 34920
tatcatggag gcacatttaa aacctgaatg aacaatgaca cgagggttaa agcaagaggc 34980
cagccgggcc tcctgcattg tcccagacag cccggctgcg cttctgttga ctcaggaaaa 35040
gctgctggaa ggagaaagat aagcgtgagc tacagcagta ccggggtggc agcaagatgg 35100
gtgaatggac ccactctggg atcagtggcg atcagcctgc aacactgcca taggatgggc 35160
aaagaaggac aaagcaagaa tggaaatgca gtgaacacca gttggttctg gactttgaaa 35220
agcagatatt tcagcttgtt gggggcccca ggcagaagaa aacctgggat ggatgtctgc 35280
tgcccactgg tgttttcaat cctgcctgtc ctggtgatgc tttcaaagga attactctgc 35340
agttctgcag gccgaggatg ctcgcagaca ccagggtcag cagacacagt aggactacag 35400
gtgtaaaaat gattctgtga tatcaagaag acattaacag atgaaatttg aaacagctgt 35460
gggaaaactt tttaacagac cggcttccct ccaaacctag ggtggattct cttgatttct 35520
gatagccaag gatttcacca cctgaaacaa actgagctct gcaaggatta acagcagcac 35580
tgttctgcag atgaatgacc tgggagcagg actagcctct catcactcct cacaggtgag 35640
gggcagaggc ctgatgccca acctcctcca gcaccgggct gaccccgctt acagaaatgt 35700
ggggatctgc tcccagaggc cagccctgca gcagccagca ccgagagctg cagaggtgcc 35760
tctgctccag caggaaagga atcctaccgg acaggaggat aaacaacagc aacaacttct 35820
tgctatcaag gacacaaaat tatcatacac ctgtatggaa gaaactattt tataaagaaa 35880
accactgaag atattttaag aagaattctt tacaaaagaa gttcatttct taaataaaaa 35940
gtattgactg actgtgtaca ttgggctcct gatcaaacat cagcacattt taaagttact 36000
ttctactatt tctgaggcag agcaagaagt tgaagcagag cagaagcaaa gaacctgctg 36060
tgacacctcc acgtagcatt aaccctaacc agcagcagga gatggatgag aagatgcttg 36120
aagacaaact agcccttacg tctgtgaaac taacatcagt ggagcagaat gacaactgaa 36180
acccctaata aaccctcctg ctattagatg agaagagatg ctgcttgata tgcattagcc 36240
tcattcctag tccatcacaa aattaggctt cactaagcaa cactacattt taattccaac 36300
aggtcatttc ctaccttgtc aagctacatt acttgacata gaaatgatgc aataagaagt 36360
cattactcaa gaaaataaaa tcctatttat tcatataccc ttgaatgtaa tggccacgtt 36420
atgtaacttg gaaattccta tgcaatgtga catggattaa tacgtattac tctttctgta 36480
ctgtttctaa atgacatttt aaacatctcg aacacaaata caaaatgcaa actcatcaac 36540
agtttcagct tctttttaag ctagtcattc caaactacta tcctcagaca ctcaactact 36600
actcccctaa tatcgggaga tacgggacac agcaagccac actgttaatt acataatcca 36660
aagagtgaac cgaatcaaca ggttgctttt tctttcctct cttttttttt tttttttaaa 36720
tgtgatttta aattctataa aaatcatcaa aactaacaac aaagcagctg ccacttttgg 36780
tcttatttgt gtagtaacat tcaaaaagct gttttaaaga aaattgcctt gcagattaac 36840
atttctcaaa gtgtaaaaat cccctttttg tccacctcat tctttaaggt cactaaatcc 36900
agaccatctg ggtaataaag ttaccatgtg tagttcccta ttcacctaat taaaacttct 36960
gtacatctcg tatggataat ttgtctatag ctaaaaccaa ctgaagtgta aaaagcagct 37020
ccggggaaat gatgtcacta gaagcaggtt actgctggca gagctggctg tcagcatatc 37080
ggcaccagga agatcccaat ttactgttaa acacatcttt tgagccaagc aacctatttc 37140
ataataataa taaatcatat ggactttcca gtaaaatcct attcacactg aaacatcaat 37200
agtaaatatt ttaatgttaa aaaaccacac agcttcgagc ctggtattta gcactatttt 37260
tgtacagagt tggtccagtc tggtagcact acggataaaa gcacgaatct ttagctgctt 37320
ccactaaaaa cataagccgc aaaagggagc aaaccattct ctcttctctc aacccgagca 37380
agagaccaag aggactgggt tttgctagtt ttggctgtta cttatgtaaa ggcagataac 37440
gaggagtcag acgccgtttt gcgtgtttcc gcagccatta cctaaaacac acgactctgt 37500
tgccaaaaag cccttgctgc cagctattac ttttcattta caagccaggt aaggatgcac 37560
tcacacagct ttttgctctg tggggtcaaa cattctgccg aggactgggg acagatgctt 37620
cctctgtggt cccaaaggac aaggattcct gtttgatacc ttcacgtgca gttgctagca 37680
atgctcccgc aagttacgga tctgcaggaa tccctgtgag aagccagttc ctaggcactg 37740
cgcttgtaat aacagagcat cacctttcaa cacagcatct tctgtaacct gtctcctaga 37800
aataaagggc tgccttccct tctctctcac gtatggaggt acttgtaaaa aggacactac 37860
agaaaactgc taatgcttca agagattaca catcctcctt gatagtatgg tatcatcaga 37920
aaataaaagc tttcctgtaa gtttacgaac agatataaaa aaacaacact acaatttatg 37980
tcagacagta ttaggatgaa agtatagaaa aaactaacac atgcttggtt aaccaagtgc 38040
actacaaaca gaaaactact ttgtcttcta cacatctgct agatccaaat tgtaactaca 38100
gtgtcataat ttggaaatac atctaacaaa ttcaggatgg acaagcgtta ggataaaatt 38160
gcttttccca gtcaatttct actcctacac tgtagaccat gcttgagtat attctagtat 38220
ccccattgag tattaatttt ggcactgtcc acacaatacc ataatgcagg ttcaataaat 38280
agaaggaccc tacatttccc ccataaaaca gtcatgctga aactgtcttg aaatactgca 38340
tctgcaagcc tatttaaagg ctaaatcttg gaatacttat cgatatggag cagtgatggg 38400
caaagtacaa gaatgcattg taagcaagtt caattaactt ctctggcaag aatataatta 38460
ctaacagaaa gcaattgcca aatatccaac taaaatagaa ggtctagcac taatgcagag 38520
caaataacta acttctcaaa acttacaaaa actttactct aacgaaatat gtctattttt 38580
acatcacttt tcccattatt ttgctaacac attttcacgt tactttatgt ggacttaccc 38640
tgtgcatgaa attctctttt cttctagttg aagaagtttt tccctgagta attagcaagc 38700
tacatcccct ttgacagaga gatactaaca agctcctaca gtttccagta agacaaggaa 38760
tccaaagcta ctaatagcaa acaagtcatt taagcaaaaa aggattacaa aggcttgttg 38820
caagagcctg gaaactcacc catatttcag attaaggctt tctgaggagc tcttttgtaa 38880
tgagttttga ctcaaatttt ttaaaatatg catttttttt aatgaaatat tgacttaaac 38940
atgtattaat cattttttct ttggacaaac aataagtgcc agaaagaccg gacaatgatt 39000
tccaatggga aaactagaga aatggtggtt ttataacaaa acctccctcg agatctcaaa 39060
gtataaagaa aaaaaatcac aaactaagaa ggatttctac tctctggtac aaggacacca 39120
gaataagctg caaatcaagc cacagtgcag cttatttgcg agacttcatt cccaaaagta 39180
cctgtcaccc catccctagg gcacatgaag tttatttcaa tgagagctac gacacagagc 39240
ggcagagccg ccaacgtacc ttgaggacct ccatgggtac ctgcgtggcg gggggcaggc 39300
tggcggggac gctgacgttg atggcaggtg tctggctgac gggcactctc tgctgcatga 39360
actgggctgc ttgctgcttc tgctgctgct cccgacgctc ctgctcctgc atctgctcac 39420
gcatgagctg ctgccgtagc aggattcgtg aggtcat 39457
<210> 2
<211> 22
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 2
tccccattct ccaaaggtta gc 22
<210> 3
<211> 22
<212> DNA
<213> 人工序列(Artificial Sequence)
<400> 3
tccccgttgt ccccacttat ta 22
Claims (2)
1. 一种采用SNP分子标记技术鉴别润州凤头白鸭黑色喙的方法,其特征是,所述分子标记位于润州凤头鸭基因组的11号染色体上,如SEQ ID NO.1所示;所述分子标记位于第28397位点,突变碱基为A或T;鉴别时,包括以下步骤:
步骤1)、提取鸭的基因组DNA;
步骤2)、针对MITF-28397设计特异性引物SEQ ID NO.2和SEQ ID NO.3进行PCR扩增:
步骤3)、将步骤2)得到的PCR产物电泳检测,然后测定基因序列,通过基因测序序列和峰图判定鸭喙色基因型;
若基因型为AA,表示受测个体为黑色喙纯合野生型基因型;若基因型为TT,表示受测个体为黄色喙纯合基因型。
2.根据权利要求1所述的方法,其特征是,步骤1)中待测鸭个体生物检材的基因组DNA为鸭翅下静脉血样基因组DNA。
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