Rice GBSS project

# Rice GBSS project # JOB https://www.nature.com/naturecareers/jobs/search?location=&text=Postdoctoral%20Fellow&facets[jobType]=Postdoctoral https://arabidopsis.org/news/jobs.jsp https://jobs.plantae.org/jobs https://jobs.sciencecareers.org/jobs/ https://recruiting.paylocity.com/recruiting/jobs/All/6b69bee1-3ec6-4d49-9564-72dfee0922ef/Boyce-Thompson-Institute-for-Plant-Research https://www.danforthcenter.org/careers/ https://buell-lab.github.io/jobs.html ## Aim 1. Identify the divergent in Wx locus using GFM algorithm. 2. Find uncovered genetic materials in genome. 3. Provide better understanding of Amylose contents in rice. ## TO DO 1. Build comprehensive and extended version of rice genome reference for SNPs and indels alignment. 2. Create haplotype genome sequence data for 3K project 3. Identify the sequence variantion in 3K rices. ## Manuscript Link https://docs.google.com/document/d/1iKYjKW4NdHkI64Zw0Bas3-jhjq7KugJ_OKnEe8u3w1A/edit?usp=sharing ## VG tube map There's more snps and indels than expected. ![](https://i.imgur.com/a7c4Cce.png) ![](https://i.imgur.com/p75qFTQ.png) ![](https://i.imgur.com/92yrewF.png) ![](https://i.imgur.com/LtNt2BH.png) ## GBSS1 locus (wx locus) in Phytozome https://phytozome-next.jgi.doe.gov/jbrowse/index.html?data=genomes%2FOsativa_v7_0&loc=Chr6%3A1765307..1767797&tracks=MSU_Rice_TE%2CTranscripts%2CAlt_Transcripts%2CPASA_assembly%2CBlastx_protein%2CBlatx_Grass%2CRepeatMasker%2CBlatx_BasalEmbryophyte%2CBlatx_BasalMonocot&highlight= ## Gene ID LOC_Os06g04200 ## Repeat ID ORSgTEMT01100006 ORSiTEMT01100003 ## MSU database http://rice.uga.edu/pub/data/Eukaryotic_Projects/o_sativa/annotation_dbs/pseudomolecules/version_7.0/chr06.dir/ ## box for Dodam, Hwayeong fastq https://ucdavis.box.com/s/cctzv4q7u50iac9drqtr1yxvqk8saomt ## Reference - Hisat-genotype https://www.nature.com/articles/s41587-019-0201-4 https://daehwankimlab.github.io/hisat-genotype/ - Shim et al., Amylose content https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9389286/ - GBSS1 paper https://www.frontiersin.org/articles/10.3389/fpls.2021.707237/full - Wx locus from 3k snps https://onlinelibrary.wiley.com/doi/10.1111/jipb.13011 - RiceVarMap2 https://ricevarmap.ncpgr.cn/ - 3K project https://gigascience.biomedcentral.com/articles/10.1186/2047-217X-3-7 https://github.com/awslabs/open-data-docs/tree/main/docs/3kricegenome - Rice pan-genome browser https://academic.oup.com/nar/article/45/2/597/2333876 - Transcript accumulation and utilization of alternate and non-consensus splice sites in rice granule-bound starch synthase are temperature-sensitive and controlled by a single-nucleotide polymorphism https://idp.springer.com/authorize/casa?redirect_uri=https://link.springer.com/article/10.1023/A:1006298608408&casa_token=cXBuYzEh34gAAAAA:DXbfdmmp8FSCbkLh35MYVj8qCPHXtnj2KaSWKlFc4B7SUDIZgexEumrNilUJ6BYixVTJBEQHVJWA43mY - The amylose content in rice endosperm is related to the post-transcriptional regulation of the waxy gene https://onlinelibrary.wiley.com/doi/abs/10.1046/j.1365-313X.1995.7040613.x ## TASSEL5 ```bash git clone https://bitbucket.org/tasseladmin/tassel-5-standalone.git cd tassel-5-standalone/ ll git pull cp ~/Downloads/248Entries_40840SNPs_inorder_21May2015_v2.txt . perl run_pipeline.pl -Xmx5g -fork1 -h 248Entries_40840SNPs_inorder_21May2015_v2.txt -export -exportType VCF -runfork1 vi 248Entries_40840SNPs_inorder_21May2015_v2.txt ``` ## VCF and HAPMAP manual https://statgen-esalq.github.io/Hapmap-and-VCF-formats-and-its-integration-with-onemap/ ## Plink2 conversion ````bash plink2 --bfile Nipponbare_indel --recode vcf --out indel --ref-from-fa Osativa_323_v7.0.fa plink2 --bfile NB_final_snp --recode vcf --out IRRI_snps --ref-from-fa Osativa_323_v7.0.fa ```` ## bcftools vcf handling ```bash bcftools concat --allow-overlaps --remove-duplicates indel.vcf IRRI_snps.vcf bcftools query -l indel.vcf bcftools query -l IRRI_snps.vcf bcftools query -l IRRI_snps.vcf > samples.txt bcftools view -S samples.txt -o filtered_indel.vcf indel.vcf bcftools concat --allow-overlaps --remove-duplicates filtered_indel.vcf IRRI_snps.vcf bcftools query -l IRRI_snps.vcf | wc -l sbatch -c 24 --mem=180g --wrap="bcftools view -S samples.txt -o IRRI_snps_filtered.vcf --threads 24 IRRI_snps.vcf" his bcf ``` ### vcf compression ```bash bgzip filtered_indel.vcf -@ 24 bgzip IRRI_snps_filtered.vcf -@ 24 ``` ## hisat-genotype ```bash git clone https://github.com/DaehwanKimLab/hisat-genotype cd hisat-genotype git submodule init git submodule update bash setup.sh ``` >LOC_Os06g04200 genomic|starch synthase, putative, expressed ACCATTCCTTCAGTTCTTTGTCTATCTCAAGACACAAATAACTGCAGTCTCTCTCTCTCT CTCTCTCTCTCTCTCTCTCTCTCTGCTTCACTTCTCTGCTTGTGTTGTTCTGTTGTTCAT CAGGAAGAACATCTGCAAGTTATACATATATGTTTATAATTCTTTGTTTCCCCTCTTATT CAGATCGATCACATGCATCTTTCATTGCTCGTTTTTCCTTACAAGTAGTCTCATACATGC TAATTTCTGTAAGGTGTTGGGCTGGAAATTAATTAATTAATTAATTGACTTGCCAAGATC CATATATATGTCCTGATATTAAATCTTCGTTCGTTATGTTTGGTTAGGCTGATCAATGTT ATTCTAGAGTCTAGAGAAACACACCCAGGGGTTTTCCAACTAGCTCCACAAGATGGTGGG CTAGCTGACCTAGATTTGAAGTCTCACTCCTTATAATTATTTTATATTAGATCATTTTCT AATATTCGTGTCTTTTTTTATTCTAGAGTCTAGATCTTGTGTTCAACTCTCGTTAAATCA TGTCTCTCGCCACTGGAGAAACAGATCAGGAGGGTTTATTTTGGGTATAGGTCAAAGCTA AGATTGAAATTCACAAATAGTAAAATCAGAATCCAACCAATTTTAGTAGCCGAGTTGGTC AAAGGAAAATGTATATAGCTAGATTTATTGTTTTGGCAAAAAAAAATCTGAATATGCAAA ATACTTGTATATCTTTGTATTAAGAAGATGAAAATAAGTAGCAGAAAATTAAAAAATGGA TTATATTTCCTGGGCTAAAAGAATTGTTGATTTGGCACAATTAAATTCAGTGTCAAGGTT TTGTGCAAGAATTCAGTGTGAAGGAATAGATTCTCTTCAAAACAATTTAATCATTCATCT GATCTGCTCAAAGCTCTGTGCATCTCCGGGTGCAACGGCCAGGATATTTATTGTGCAGTA AAAAAATGTCATATCCCCTAGCCACCCAAGAAACTGCTCCTTAAGTCCTTATAAGCACAT ATGGCATTGTAATATATATGTTTGAGTTTTAGCGACAATTTTTTTAAAAACTTTTGGTCC TTTTTATGAACGTTTTAAGTTTCACTGTCTTTTTTTTTCGAATTTTAAATGTAGCTTCAA ATTCTAATCCCCAATCCAAATTGTAATAAACTTCAATTCTCCTAATTAACATCTTAATTC ATTTATTTGAAAACCAGTTCAAATTCTTTTAGGCTCACCAAACCTTAAACAATTCAATTC AGTGCAGAGATCTTCCACAGCAACAGCTAGACAACCACCATGTCGGCTCTCACCACGTCC CAGCTCGCCACCTCGGCCACCGGCTTCGGCATCGCCGACAGGTCGGCGCCGTCGTCGCTG CTCCGCCACGGGTTCCAGGGCCTCAAGCCCCGCAGCCCCGCCGGCGGCGACGCGACGTCG CTCAGCGTGACGACCAGCGCGCGCGCGACGCCCAAGCAGCAGCGGTCGGTGCAGCGTGGC AGCCGGAGGTTCCCCTCCGTCGTCGTGTACGCCACCGGCGCCGGCATGAACGTCGTGTTC GTCGGCGCCGAGATGGCCCCCTGGAGCAAGACCGGCGGCCTCGGTGACGTCCTCGGTGGC CTCCCCCCTGCCATGGCTGTAAGCACACACAAACTTCGATCGCTCGTCGTCGCTGACCGT CGTCGTCTTCAACTGTTCTTGATCATCGCATTGGATGGATGTGTAATGTTGTGTTCTTGT GTTCTTTGCAGGCGAATGGCCACAGGGTCATGGTGATCTCTCCTCGGTACGACCAGTACA AGGACGCTTGGGATACCAGCGTTGTGGCTGAGGTAGGAGCATATGCGTGATCAGATCATC ACAAGATCGATTAGCTTTAGATGATTTGTTACATTTCGCAAGATTTTAACCCAAGTTTTT GTGGTGCAATTCATTGCAGATCAAGGTTGCAGACAGGTACGAGAGGGTGAGGTTTTTCCA TTGCTACAAGCGTGGAGTCGACCGTGTGTTCATCGACCATCCGTCATTCCTGGAGAAGGT GGAGTCATCATTAGTTTACCTTTTTTGTTTTTACTGAATTATTAACAGTGCATTTAGCAG TTGGACTGAGCTTAGCTTCCACTGGTGATTTCAGGTTTGGGGAAAGACCGGTGAGAAGAT CTACGGACCTGACACTGGAGTTGATTACAAAGACAACCAGATGCGTTTCAGCCTTCTTTG CCAGGTCAGTGATTACTTCTATCTGATGATGGTTGGAAGCATCACGAGTTTACCATAGTA TGTATGGATTCATAACTAATTCGTGTATTGATGCTACCTGCAGGCAGCACTCGAGGCTCC TAGGATCCTAAACCTCAACAACAACCCATACTTCAAAGGAACTTATGGTGAGTTACAATT GATCTCAAGATCTTATAACTTTCTTCGAAGGAATCCATGATGATCAGACTAATTCCTTCC GGTTTGTTACTGACAACAGGTGAGGATGTTGTGTTCGTCTGCAACGACTGGCACACTGGC CCACTGGCGAGCTACCTGAAGAACAACTACCAGCCCAATGGCATCTACAGGAATGCAAAG GTCTATGCTTGTTCTTGCCATACCAACTCAAATCTGCATGCACACTGCATTCTGTTCAGA AACTGACTGTCTGAATCTTTTTCACTGCAGGTTGCTTTCTGCATCCACAACATCTCCTAC CAGGGCCGTTTCGCTTTCGAGGATTACCCTGAGCTGAACCTCTCCGAGAGGTTCAGGTCA TCCTTCGATTTCATCGACGGGTATGAGTAAGATTCTAAGAGTAACTTACTGTCAATTCGC CATATATCGATTCAATCCAAGATCCTTTTGAGCTGACAACCCTGCACTACTGTCCATCGT TCAAATCCGGTTAAATTTCAGGTATGACACGCCGGTGGAGGGCAGGAAGATCAACTGGAT GAAGGCCGGAATCCTGGAAGCCGACAGGGTGCTCACCGTGAGCCCGTACTACGCCGAGGA GCTCATCTCCGGCATCGCCAGGGGATGCGAGCTCGACAACATCATGCGGCTCACCGGCAT CACCGGCATCGTCAACGGCATGGACGTCAGCGAGTGGGATCCTAGCAAGGACAAGTACAT CACCGCCAAGTACGACGCAACCACGGTAAGAACGAATGCATTCTTCACAAGATATGCAAT CTGAATTTTCTTTGAAAAAGAAATTATCATCTGTCACTTCTTGATTGATTCTGACAAGGC AAGAATGAGTGACAAATTTCAGGCAATCGAGGCGAAGGCGCTGAACAAGGAGGCGTTGCA GGCGGAGGCGGGTCTTCCGGTCGACAGGAAAATCCCACTGATCGCGTTCATCGGCAGGCT GGAGGAACAGAAGGGCCCTGACGTCATGGCCGCCGCCATCCCGGAGCTCATGCAGGAGGA CGTCCAGATCGTTCTTCTGGTATAATATAATACACTACAAGACACACTTGCACGATATGC CAAAAATTCAGAACAAATTCAGTGGCAAAAAAAAAACTCGAATATTAGGGAAGGACCTAA TAATATCAAATAATTAGAAGGGGTGAGGCTTTGAACCCAGATCGTCTAGTCCACCACCTT GTGGAGTTAGCCGGAAGACCTCTGAGCATTTCTCAATTCAGTGGCAAATGATGTGTATAA TTTTGATCCGTGTGTGTTTCAGGGTACTGGAAAGAAGAAGTTCGAGAAGCTGCTCAAGAG CATGGAGGAGAAGTATCCGGGCAAGGTGAGGGCCGTGGTGAAGTTCAACGCGCCGCTTGC TCATCTCATCATGGCCGGAGCCGACGTGCTCGCCGTCCCCAGCCGCTTCGAGCCCTGTGG ACTCATCCAGCTGCAGGGGATGAGATACGGAACGGTATACAATTTCCATCTATCAATTCG ATTGTTCGATTTCATCTTTGTGCAATGCAATGCAATTGCAAATGCAAATGCATGATGATT TTCCTTGTTGATTTCTCCAGCCCTGTGCTTGCGCGTCCACCGGTGGGCTCGTGGACACGG TCATCGAAGGCAAGACTGGTTTCCACATGGGCCGTCTCAGCGTCGACGTAAGCCTATACA TTTACATAACAATCAGATATGACACATCCTAATACCGATAAGTCGGTACACTACTACACA TTTACATGGTTGCTGGTTATATGGTTTTTTTGGCAGTGCAAGGTGGTGGAGCCAAGCGAC GTGAAGAAGGTGGCGGCCACCCTGAAGCGCGCCATCAAGGTCGTCGGCACGCCGGCGTAC GAGGAGATGGTCAGGAACTGCATGAACCAGGACCTCTCCTGGAAGGTATAAATTACGAAA CAAATTTAACCCAAACATATACTATATACTCCCTCCGCTTCTAAATATTCAACGCCGTTG TCTTTTTTAAATATGTTTGACCATTCGTCTTATTAAAAAAATTAAATAATTATAAATTCT TTTCCTATCATTTGATTCATTGTTAAATATACTTATATGTATACATATAGTTTTACATAT TTCATAAAATTTTTTGAACAAGACGAACGGTCAAACATGTGCTAAAAAGTTAACGGTGTC GAATATTCAGAAACGGAGGGAGTATAAACGTCTTGTTCAGAAGTTCAGAGATTCACCTGT CTGATGCTGATGATGATTAATTGTTTGCAACATGGATTTCAGGGGCCTGCGAAGAACTGG GAGAATGTGCTCCTGGGCCTGGGCGTCGCCGGCAGCGCGCCGGGGATCGAAGGCGACGAG ATCGCGCCGCTCGCCAAGGAGAACGTGGCTGCTCCTTGAAGAGCCTGAGATCTACATATG GAGTGATTAATTAATATAGCAGTATATGGATGAGAGACGAATGAACCAGTGGTTTGTTTG TTGTAGTGAATTTGTAGCTATAGCCAATTATATAGGCTAATAAGTTTGATGTTGTACTCT TCTGGGTGTGCTTAAGTATCTTATCGGACCCTGAATTTATGTGTGTGGCTTATTGCCAAT AATATTAAGTAATAAAGGGTTTATTATATTATTATATATGTTATATTATACTTCC >LOC_Os06g04200.1 protein|starch synthase, putative, expressed MSALTTSQLATSATGFGIADRSAPSSLLRHGFQGLKPRSPAGGDATSLSVTTSARATPKQ QRSVQRGSRRFPSVVVYATGAGMNVVFVGAEMAPWSKTGGLGDVLGGLPPAMAANGHRVM VISPRYDQYKDAWDTSVVAEIKVADRYERVRFFHCYKRGVDRVFIDHPSFLEKVWGKTGE KIYGPDTGVDYKDNQMRFSLLCQAALEAPRILNLNNNPYFKGTYGEDVVFVCNDWHTGPL ASYLKNNYQPNGIYRNAKVAFCIHNISYQGRFAFEDYPELNLSERFRSSFDFIDGYDTPV EGRKINWMKAGILEADRVLTVSPYYAEELISGIARGCELDNIMRLTGITGIVNGMDVSEW DPSKDKYITAKYDATTAIEAKALNKEALQAEAGLPVDRKIPLIAFIGRLEEQKGPDVMAA AIPELMQEDVQIVLLGTGKKKFEKLLKSMEEKYPGKVRAVVKFNAPLAHLIMAGADVLAV PSRFEPCGLIQLQGMRYGTPCACASTGGLVDTVIEGKTGFHMGRLSVDCKVVEPSDVKKV AATLKRAIKVVGTPAYEEMVRNCMNQDLSWKGPAKNWENVLLGLGVAGSAPGIEGDEIAP LAKENVAAP* >LOC_Os06g04200.2 protein|starch synthase, putative, expressed MSALTTSQLATSATGFGIADRSAPSSLLRHGFQGLKPRSPAGGDATSLSVTTSARATPKQ QRSVQRGSRRFPSVVVYATGAGMNVVFVGAEMAPWSKTGGLGDVLGGLPPAMAANGHRVM VISPRYDQYKDAWDTSVVAEIKVADRYERVRFFHCYKRGVDRVFIDHPSFLEKVWGKTGE KIYGPDTGVDYKDNQMRFSLLCQAALEAPRILNLNNNPYFKGTYGEDVVFVCNDWHTGPL ASYLKNNYQPNGIYRNAKVAFCIHNISYQGRFAFEDYPELNLSERFRSSFDFIDGYDTPV EGRKINWMKAGILEADRVLTVSPYYAEELISGIARGCELDNIMRLTGITGIVNGMDVSEW DPSKDKYITAKYDATTAIEAKALNKEALQAEAGLPVDRKIPLIAFIGRLEEQKGPDVMAA AIPELMQEDVQIVLLGTGKKKFEKLLKSMEEKYPGKVRAVVKFNAPLAHLIMAGADVLAV PSRFEPCGLIQLQGMRYGTPCACASTGGLVDTVIEGKTGFHMGRLSVDCKVVEPSDVKKV AATLKRAIKVVGTPAYEEMVRNCMNQDLSWKGPAKNWENVLLGLGVAGSAPGIEGDEIAP LAKENVAAP* >LOC_Os06g04200.4 protein|starch synthase, putative, expressed MSALTTSQLATSATGFGIADRSAPSSLLRHGFQGLKPRSPAGGDATSLSVTTSARATPKQ QRSVQRGSRRFPSVVVYATGAGMNVVFVGAEMAPWSKTGGLGDVLGGLPPAMAANGHRVM VISPRYDQYKDAWDTSVVAEIKVADRYERVRFFHCYKRGVDRVFIDHPSFLEKVWGKTGE KIYGPDTGVDYKDNQMRFSLLCQAALEAPRILNLNNNPYFKGTYGEDVVFVCNDWHTGPL ASYLKNNYQPNGIYRNAKVAFCIHNISYQGRFAFEDYPELNLSERFRSSFDFIDGYDTPV EGRKINWMKAGILEADRVLTVSPYYAEELISGIARGCELDNIMRLTGITGIVNGMDVSEW DPSKDKYITAKYDATTAIEAKALNKEALQAEAGLPVDRKIPLIAFIGRLEEQKGPDVMAA AIPELMQEDVQIVLLGTGKKKFEKLLKSMEEKYPGKVRAVVKFNAPLAHLIMAGADVLAV PSRFEPCGLIQLQGMRYGTPCACASTGGLVDTVIEGKTGFHMGRLSVDCKVVEPSDVKKV AATLKRAIKVVGTPAYEEMVRNCMNQDLSWKGPAKNWENVLLGLGVAGSAPGIEGDEIAP LAKENVAAP* >LOC_Os06g04200.3 protein|starch synthase, putative, expressed MSALTTSQLATSATGFGIADRSAPSSLLRHGFQGLKPRSPAGGDATSLSVTTSARATPKQ QRSVQRGSRRFPSVVVYATGAGMNVVFVGAEMAPWSKTGGLGDVLGGLPPAMAANGHRVM VISPRYDQYKDAWDTSVVAEIKVADRYERVRFFHCYKRGVDRVFIDHPSFLEKVWGKTGE KIYGPDTGVDYKDNQMRFSLLCQAALEAPRILNLNNNPYFKGTYGEDVVFVCNDWHTGPL ASYLKNNYQPNGIYRNAKVAFCIHNISYQGRFAFEDYPELNLSERFRSSFDFIDGYDTPV EGRKINWMKAGILEADRVLTVSPYYAEELISGIARGCELDNIMRLTGITGIVNGMDVSEW DPSKDKYITAKYDATTAIEAKALNKEALQAEAGLPVDRKIPLIAFIGRLEEQKGPDVMAA AIPELMQEDVQIVLLGTGKKKFEKLLKSMEEKYPGKVRAVVKFNAPLAHLIMAGADVLAV PSRFEPCGLIQLQGMRYGTPCACASTGGLVDTVIEGKTGFHMGRLSVDCKVVEPSDVKKV AATLKRAIKVVGTPAYEEMVRNCMNQDLSWKGPAKNWENVLLGLGVAGSAPGIEGDEIAP LAKENVAAP* >FGENESH MSALTTSQLATSATGFGIADRSAPSSLLRHGFQGLKPRSPAGGDATSLSVTTSARATPKQ QRSVQRGSRRFPSVVVYATGAGMNVVFVGAEMAPWSKTGGLGDVLGGLPPAMAANGHRVM VISPRYDQYKDAWDTSVVAEIKVADRYERVRFFHCYKRGVDRVFIDHPSFLEKVWGKTGE KIYGPDTGVDYKDNQMRFSLLCQAALEAPRILNLNNNPYFKGTYGEDVVFVCNDWHTGPL ASYLKNNYQPNGIYRNAKVAFCIHNISYQGRFAFEDYPELNLSERFRSSFDFIDGYDTPV EGRKINWMKAGILEADRVLTVSPYYAEELISGIARGCELDNIMRLTGITGIVNGMDVSEW DPSKDKYITAKYDATTARMSDKFQAIEAKALNKEALQAEAGLPVDRKIPLIAFIGRLEEQ KGPDVMAAAIPELMQEDVQIVLLGTGKKKFEKLLKSMEEKYPGKVRAVVKFNAPLAHLIM AGADVLAVPSRFEPCGLIQLQGMRYGTPCACASTGGLVDTVIEGKTGFHMGRLSVDCKVV EPSDVKKVAATLKRAIKVVGTPAYEEMVRNCMNQDLSWKGPAKNWENVLLGLGVAGSAPG IEGDEIAPLAKENVAAP https://phytozome-next.jgi.doe.gov/jbrowse/index.html?data=genomes%2FOsativa_v7_0&loc=Chr6%3A1765267..1767757&tracks=MSU_Rice_TE%2CTranscripts%2CAlt_Transcripts%2CPASA_assembly%2CBlastx_protein%2CBlatx_Grass%2CRepeatMasker&highlight= ## KC ### 2022.12.21 -Wxb (Japonica) transcript include splicing form and non-splicing form (including intron1), and Wxa (Indica) transcript only has splicing form. ![](https://i.imgur.com/IAofnQe.png) - Varisous splicing variants were observed in Wx ![](https://i.imgur.com/AjFvRjC.png) ![](https://i.imgur.com/0aaAeie.png) - Repeat elements detected from RepeatMasker https://phytozome-next.jgi.doe.gov/jbrowse/index.html?data=genomes%2FOsativa_v7_0&loc=Chr6%3A1765958..1766450&tracks=MSU_Rice_TE%2CPASA_assembly%2CBlastx_protein%2CBlatx_Grass%2CTranscripts%2CAlt_Transcripts%2CRepeatMasker&highlight= ![](https://i.imgur.com/goxbO14.png) - ![](https://i.imgur.com/NO17V9g.png) ### 2022.12.28 In the literature search, three characteristics of rice Waxy gene were found. 1. Canonical splicing - T/G SNP at 1,765,761 on chromosome 6 regulates alternative splicing variant. Japonica rice which has low amylose content harbor T-type SNP and T-type varieties showed two types of Wx transcript forms 3.3kb and 2.3kb. 2.3kb transcript is matured transcript without intron, while 3.3kb transcript includes 1st intron. The waxy mutant which has no amylose in the endosperm also have about 3.3kb transcript but Wx protein was not detected, indicating that 3.3kb transcript may not be translated. On the other hands, G-type SNP is mostly found in indica rice and displayed high amylose content. G-type varieties have only 2.3kb of matured transcript form. - In addition, aberrant splicing of intron 1 was identified and more than six splicing patterns were observed (Cai et al. 1998; Larkin and Park 1999). 2. Wx gene is temperature sensitive and controlled by T/G SNP - T-type varieties showed decreased gene expression of 2.3kb and 3.3kb at the high temperature, leading decreased amylose content. However, G-type varieties showed consistent expression level at low and high temperature. 3. Repeat sequence in 1 intron. - Two SINE elements were identified on 5’ terminal of Wx gene (ORSiTEMT01100003 and ORSgTEMT01100006). RNA secondary structure is predicted with 5’ terminal region (350bp) of Wx gene and it formed six cruciforms (Cai et al. 1998). Q&A from last discussion 1. Discussions about first intron repeat. - Many studies have been conducted about first intron repeat of Wx gene in rice. Wang et al. (1994) reported that two types of transposon-like elements (RTL-1 and RTL-2) were located in introns 1, 10, and 13 of the rice Wx gene. In addition, Wang et al. (1995) predicted RNA secondary structure in the 5’ terminal region (350 bp) of Wx gene. 2. Rice 3k database in amylose content data. - Rice 3k database include amylose content data and ENDO (endosperm type) are evaluated by 1 (Non-waxy), 2 (Waxy), 3 (Indeterminate), and 999 (Mixture). - IRRI group published this paper <Haplotype analysis of key genes governing grain yield and quality traits across 3K RG panel reveals scope for the development of tailor-made rice with enhanced genetic gains, Plant biotechnology journal, doi: 10.1111/pbi.13087> and they measured amylose content. But amylose content data is not included in supplementary files. 3. Do transposable elements have structure? Yse. RCSB PDB has LINE or SINE structures. RNA hairpin or RNA stem-loop structures are experimentally observed. Also, AlphaFold2 predict computed structure model of retrotransposon proteins. 4. Rice 3K database has RNA-seq data? - I think Rice 3K database doesn’t have RNA-seq data. Rice pan-genome browser was constructed by rice3k db and they used public RNA-seq data. The sample IDs were listed in supplementary table1. 5. GBSS1 has four different types of transcript form. Do they have different expression level and SINE is associated with expression level? - Expressional level of LOC_Os06g04200.1-4 was not examined from previous studies. SINE in intron 1 possibly associated with Wx expression level. 01/18/2023 Please check below manuscript https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7663775/ Main questions are 1. does isoform variation affect to starch contents? - Wxb type expressed two major types of transcripts 2.3 and 3.3 kb forms. The 2.3 Kb form is completely matured type and 3.3 Kb is prematured form which has 1 Kb of intron 1 and is not normally function. 2. Above manuscript includes, aus, indica and jap, do they have difference splicing form in GBSS1? - Above manuscript opened only N22 (variety name) transcriptome data. Three transcript forms were identified and one of them showed different transcript form compared to reference isoforms. (I found their transcriptome data from NCBI GSE153030, https://www.ncbi.nlm.nih.gov/geo/download/?acc=GSE153030&format=file&file=GSE153030%5Ffinal%5FdeNovo%2EB%5FN22%2Efasta%2Egz) 3. Can we interpret the splicing form from their work? - 4. Do they have divergent starch contents? - They didn't measure the amylose content, but they have different amylose content due to their different ecotype. 5. do they have DNA-seq data in 3k? - Only one variety (Dular) was included in rice 3k. - However, other varieties are popular, so we may be able to find sequencing data from NCBI. https://github.com/pangenome/pggb 03/07/23 - Table. Rice 3K DB endosperm type ![](https://i.imgur.com/tQoVkYA.png) - Manhattan plot generated from Rice 3K DB using endosperm type phenotype ![](https://i.imgur.com/wnkMkwx.png) https://pubmed.ncbi.nlm.nih.gov/34051138/

Syntax	Example	Reference
# Header	Header	基本排版
- Unordered List	Unordered List
1. Ordered List	Ordered List
- [ ] Todo List	Todo List
> Blockquote	Blockquote
Bold font	Bold font
Italics font	Italics font
~~Strikethrough~~	~~Strikethrough~~
19^th^	19^th
H~2~O	H₂O
++Inserted text++	Inserted text
==Marked text==	Marked text
[link text](https:// "title")	Link
![image alt](https:// "title")	Image
`Code`	`Code`	在筆記中貼入程式碼
```javascript var i = 0; ```	`var i = 0;`
:smile:		Emoji list
{%youtube youtube_id %}	Externals
$L^aT_eX$	L^aT_eX
:::info This is a alert area. :::	This is a alert area.